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An important advantage of computed tomography coronary angiography (CCTA) is its ability to visualize the presence and severity of atherosclerotic plaque, rather than just assessing coronary artery stenoses. Until recently, assessment of plaque subtypes on CCTA relied on visual assessment of the extent of calcified/non-calcified plaque, or visually identifying high-risk plaque characteristics. Recent software developments facilitate the quantitative assessment of plaque volume or burden on CCTA, and the identification of subtypes of plaque based on their attenuation density. These techniques have shown promise in single and multicenter studies, demonstrating that the amount and type of plaque are associated with subsequent cardiac events. However, there are a number of limitations to the application of these techniques, including the limitations imposed by the spatial resolution of current CT scanners, challenges from variations between reconstruction algorithms, and the additional time to perform these assessments. At present, these are a valuable research technique, but not yet part of routine clinical practice. Future advances that improve CT resolution, standardize acquisition techniques and reconstruction algorithms and automate image analysis will improve the clinical utility of these techniques. This review will discuss the technical aspects of quantitative plaque analysis and present pro and con arguments for the routine use of quantitative plaque analysis on CCTA., Competing Interests: Declaration of competing interest JPE Equity and Consultant, Cleerly Healthcare Inc. The other authors have no conflicts of interest., (Copyright © 2021 Society of Cardiovascular Computed Tomography. Published by Elsevier Inc. All rights reserved.)

Coronary computed tomographic angiography (CCTA) provides a wealth of clinically meaningful information beyond anatomic stenosis alone, including the presence or absence of nonobstructive atherosclerosis and high-risk plaque features as precursors for incident coronary events. There is, however, no uniform agreement on how to identify and quantify these features or their use in evidence-based clinical decision-making. This statement from the Society of Cardiovascular Computed Tomography and North American Society of Cardiovascular Imaging addresses this gap and provides a comprehensive review of the available evidence on imaging of coronary atherosclerosis. In this statement, we provide standardized definitions for high-risk plaque (HRP) features and distill the evidence on the effectiveness of risk stratification into usable practice points. This statement outlines how this information should be communicated to referring physicians and patients by identifying critical elements to include in a structured CCTA report - the presence and severity of atherosclerotic plaque (descriptive statements, CAD-RADS™ categories), the segment involvement score, HRP features (e.g., low attenuation plaque, positive remodeling), and the coronary artery calcium score (when performed). Rigorous documentation of atherosclerosis on CCTA provides a vital opportunity to make recommendations for preventive care and to initiate and guide an effective care strategy for at-risk patients., (Copyright © 2020. Published by Elsevier Inc.)

The purpose of this review is to summarize the work published by the Journal of Cardiovascular Computed Tomography (JCCT) for the year 2019, highlighting original research and new guidelines., Competing Interests: Declaration of competing interest CNC receives research support from and has received speaker fees from Siemens Healthineers. JAL serves as a consultant and holds stock options with Circle CVI and HeartFlow and receives research support from GE Healthcare. JKM receives funding from the Dalio Foundation, NIH, and GE Healthcare. JKM also serves on the scientific advisory board of Arineta and GE Healthcare, and has an equity interest in Cleerly., (Copyright © 2020 Society of Cardiovascular Computed Tomography. Published by Elsevier Inc. All rights reserved.)

This expert consensus statement from the Society of Cardiovascular Computed Tomography (SCCT) provides an evidence synthesis on the use of computed tomography (CT) imaging for diagnosis and risk stratification of coronary artery disease in women. From large patient and population cohorts of asymptomatic women, detection of any coronary artery calcium that identifies females with a 10-year atherosclerotic cardiovascular disease risk of >7.5% may more effectively triage women who may benefit from pharmacologic therapy. In addition to accurate detection of obstructive coronary artery disease (CAD), CT angiography (CTA) identifies nonobstructive atherosclerotic plaque extent and composition which is otherwise not detected by alternative stress testing modalities. Moreover, CTA has superior risk stratification when compared to stress testing in symptomatic women with stable chest pain (or equivalent) symptoms. For the evaluation of symptomatic women both in the emergency department and the outpatient setting, there is abundant evidence from large observational registries and multi-center randomized trials, that CT imaging is an effective procedure. Although radiation doses are far less for CT when compared to nuclear imaging, radiation dose reduction strategies should be applied in all women undergoing CT imaging. Effective and appropriate use of CT imaging can provide the means for improved detection of at-risk women and thereby focus preventive management resulting in long-term risk reduction and improved clinical outcomes., (Copyright © 2018 Society of Cardiovascular Computed Tomography. All rights reserved.)

Evaluation of the left atrium and left atrial appendage for the presence of thrombus prior to cardioversion and pulmonary vein isolation, and of the entire heart for embolic sources in the setting of cryptogenic stroke, has long been standard medical care. Guidelines have uniformly recommended transesophageal echocardiography (TEE) to accomplish these goals. In recent years, computed tomographic angiography has demonstrated diagnostic accuracy similar to that of TEE for the detection of thrombus. Analysis of the pertinent data and relative merits of the 2 technologies leads to the conclusions that: 1) both modalities have some unique, nonoverlapping capabilities that may dictate their use in specific situations; 2) computed tomographic angiography is a reasonable alternative to TEE when the primary aim is to exclude left atrial and left atrial appendage thrombus and in patients in whom the risks associated with TEE outweigh the benefits; and 3) both options should be discussed with the patient in the setting of shared decision making., (Copyright © 2018 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

The rising cost of healthcare is prompting numerous policy and advocacy discussions regarding strategies for constraining growth and creating a more efficient and effective healthcare system. Cardiovascular imaging is central to the care of patients at risk of, and living with, heart disease. Estimates are that utilization of cardiovascular imaging exceeds 20 million studies per year. The Society of Cardiovascular CT (SCCT), alongside Rush University Medical Center, and in collaboration with government agencies, regional payers, and industry healthcare experts met in November 2016 in Chicago, IL to evaluate obstacles and hurdles facing the cardiovascular imaging community and how they can contribute to efficacy while maintaining or even improving outcomes and quality. The summit incorporated inputs from payers, providers, and patients' perspectives, providing a platform for all voices to be heard, allowing for a constructive dialogue with potential solutions moving forward. This article outlines the proceedings from the summit, with a detailed review of past hurdles, current status, and potential solutions as we move forward in an ever-changing healthcare landscape., (Copyright © 2017 Society of Cardiovascular Computed Tomography. All rights reserved.)

Objectives: The aim of this study was to assess the relationship of the extent of atherosclerosis on coronary computed tomographic angiography to the extent of secondhand tobacco smoke (SHTS) exposure in asymptomatic never smokers., Background: A dose-related association between SHTS and coronary artery calcium has been reported, but the total extent of atherosclerosis has not been investigated., Methods: A total of 268 never smokers, ages 40 to 80 years, completed a questionnaire assessing risk factors and extent of lifetime SHTS exposure, providing a total SHTS exposure score. Ordinal coronary artery calcium scores were derived from low-dose nongated computed tomographic scans, followed by computed tomographic angiography. Analyses of the prevalence, extent, and plaque characteristics of atherosclerosis were performed, and the independent contribution of SHTS, adjusted for other documented risk factors, was determined., Results: Coronary atherosclerosis was noted in 48% and was more frequent with low to moderate and high versus minimal SHTS exposure (48% and 69% vs. 25%; p < 0.0001). Adjusted odds ratios for any atherosclerosis were 2.1 (95% confidence interval: 1.0 to 4.4; p = 0.05) for low to moderate and 3.5 (95% confidence interval: 1.4 to 8.5; p = 0.01) for high exposure versus minimal SHTS exposure and were not significant for standard risk factors of diabetes (p = 0.56), hyperlipidemia (p = 0.11), hypertension (p = 0.65), and renal disease (p = 0.24). With increasing SHTS exposure, the percentage of major vessel (14%, 41%, and 45%; p = 0.0013) with any plaque or stenosis increased, as did the number with 5 or more involved segments (0%, 39%, and 61%; p = 0.0001). Also the average number of involved segments increased (0.82, 1.98, and 3.49; p < 0.0001), with calcified plaques alone (0.25, 0.77, and 1.52; p < 0.0001), with calcified and partially calcified plaques (0.28, 0.82, and 1.58; p < 0.001), but not with noncalcified plaques alone (p = 0.11)., Conclusions: The presence and extent of atherosclerosis were associated with the extent of SHTS exposure even when adjusted for other risk factors, further demonstrating the causal relationship of SHTS exposure and coronary disease., (Copyright © 2017 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Background and Objectives: Patients with social anxiety disorder suffer from pronounced fears in social situations. As gaze perception is crucial in these situations, we examined which factors influence the range of gaze directions where mutual gaze is experienced (the cone of gaze)., Methods: The social stimulus was modified by changing the number of people (heads) present and the emotional expression of their faces. Participants completed a psychophysical task, in which they had to adjust the eyes of a virtual head to gaze at the edge of the range where mutual eye-contact was experienced., Results: The number of heads affected the width of the gaze cone: the more heads, the wider the gaze cone. The emotional expression of the virtual head had no consistent effect on the width of the gaze cone, it did however affect the emotional state of the participants. Angry expressions produced the highest arousal values. Highest valence emerged from happy faces, lowest valence from angry faces., Conclusion: These results suggest that the widening of the gaze cone in social anxiety disorder is not primarily mediated by their altered emotional reactivity. Implications for gaze assessment and gaze training in therapeutic contexts are discussed., Limitations: Due to interindividual variability, enlarged gaze cones are not necessarily indicative of social anxiety disorder, they merely constitute a correlate at the group level., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

This expert consensus statement summarizes the available data regarding the prognostic value of CAC in the asymptomatic population and its ability to refine individual risk prediction, addresses the limitations identified in the current traditional risk factor-based treatment strategies recommended by the 2013 ACC/AHA Prevention guidelines including use of the Pooled Cohort Equations (PCE), and the US Preventive Services Task Force (USPSTF) Recommendation Statement for Statin Use for the Primary Prevention of Cardiovascular Disease in Adults. It provides CAC based treatment recommendations both within the context of the shared decision making model espoused by the 2013 ACC/AHA Prevention guidelines and independent of these guidelines., (Copyright © 2017 Society of Cardiovascular Computed Tomography. Published by Elsevier Inc. All rights reserved.)

Background: The incremental prognostic value of the plaque features in coronary computed tomographic angiography (CTA) has not been well assessed. This study was designed to determine whether CTA high-risk plaques have prognostic value incremental to the Framingham risk score (FRS) and the severity of luminal obstruction., Methods: A total of 628 newly symptomatic patients without known coronary artery disease underwent CTA. They were followed for a median of 677 days during which there were 26 cardiac events, including cardiac death, acute myocardial infarction, and hospitalization for unstable angina. Incremental prognostic value of adding plaque characteristics to the number of diseased vessels and the FRS was evaluated using 3 Cox models and net reclassification indexes., Results: The discrimination index was significantly increased by adding the number of diseased vessels to the FRS (change in c-statistic from 65.8% to 78.6%, p=0.028) but not significantly by further adding plaque characteristics (change in c-statistic from 78.6% to 80.0%, p=0.812). However, improved model-fitting by adding plaque characteristics into the linear combination with risk score and the number of diseased vessels (p=0.007 from likelihood ratio test) and the lowest value of Akaike's information criteria of that model indicated that plaque characteristics improved both predictive accuracy and discrimination perspective. More subjects reclassified by plaque characteristics were moved to directions consistent with their subsequent cardiac event status than in an inconsistent direction., Conclusions: Evaluation of CTA plaque characteristics may provide incremental prognostic value to the number of diseased vessels and the FRS., (Copyright © 2015 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.)

Objective: To compare Agatston with Ordinal Scores for the extent of coronary artery calcification (coronary artery calcium) using nongated low-dose computed tomography (CT) scans., Materials and Methods: A total of 631 asymptomatic participants had CT scans from 2010 to 2013. Their Ordinal and Agatston Score were classified into categories., Results: The Ordinal Score Categories showed excellent agreement (weighted kappa of 0.83; 95% confidence interval: 0.79-0.88) with the Agatston Score Categories., Conclusions: The use of the Ordinal Score is readily obtained on low-dose CT scans that are used for CT screening for lung cancer and these scores are useful for risk stratification of coronary artery disease., (Copyright © 2015. Published by Elsevier Inc.)

Background: Recent research has begun to examine the neurophysiologic basis of pathological gambling. However, direct evidence of a behavioral deficit and an accompanying neurofunctional deviation in a realistic gambling context such as Black Jack has not yet been reported., Methods: Electroencephalogram was recorded while 20 problem gamblers and 21 control participants played a computerized version of Black Jack. Participants were asked to decide at point scores between 11 and 21 whether they wanted to take another card ("hit") to arrive closer to 21 than the opponent (simulated by computer) or not to take another card ("sit") to avoid going over 21 ("bust")., Results: At a critical point score of 16, problem gamblers decided more often to hit despite losses due to a bust on the preceding trial, whereas control participants decided more often to sit under these conditions. Furthermore, problem gamblers showed more reward-related positive amplitudes in the event-related brain potential than control participants after successful hit decisions at 16., Conclusions: Here we provide experimental evidence for high-risk taking behavior in gamblers and its correlate in event-related brain potentials. Our results suggest that high-risk-taking behavior in problem gamblers is associated with an increased reward-related neural response to infrequent successes of this behavior., (Copyright 2010 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Background: Digital thermal monitoring (DTM) of vascular function was shown to correlate with the presence of known coronary artery disease (CAD)., Objective: We evaluated whether DTM can identify at-risk, asymptomatic patients with significant coronary artery calcium (CAC) or increased Framingham risk score (FRS)., Methods: Two hundred thirty-three consecutive asymptomatic subjects (58 +/- 11 years; 62% men) without known CAD underwent DTM, CAC, and FRS calculation. DTM measurements were obtained during and after a 5-minute suprasystolic arm-cuff occlusion. After cuff-deflation temperature rebound (TR) and area under the temperature curve (AUC) were measured and correlated with FRS and CAC., Results: TR was lower in patients with FRS > 20% and CAC >or= 100 as compared with FRS < 10% and CAC < 10, respectively (P < 0.05). After adjustment for age, sex, and traditional cardiac risk factors, the odds ratio of the lowest compared with the upper 2 tertiles of TR was 3.96 for FRS >or= 20% and 2.37 for CAC >or= 100 compared with low-risk cohorts. The area under the receiver operating characteristic (ROC) curve to predict CAC >or= 100 increased significantly from 0.66 for FRS to 0.79 for TR to 0.89 for TR + FRS., Conclusions: Vascular dysfunction measured by DTM strongly correlates with FRS and CAC independent of age, sex, and traditional cardiac risk factors and was superior to FRS for the prediction of significant CAC.

Objective: It is thought that emphysema patients are at a higher risk of coronary artery disease. The present study is one of very few that evaluated the prevalence of significant coronary artery disease in emphysema patients using coronary artery calcification measured by electron beam computed tomography., Methods: A retrospective chart review evaluated 1720 consecutive patients, some of whom were self-referred. All patients had both heart and lungs imaged with electron beam computed tomography when they were seen at the Inner Imaging Center, a cardiac imaging center affiliated with the Beth Israel Hospital in New York, New York. Multiple logistic regression was performed to determine which factors were independently associated with coronary artery calcification., Results: Age, sex, hypertension and smoking were the risk factors independently associated with coronary artery calcification in the population studied. The emphysema group was significantly higher on measures of smoking and hypertension compared with the control group. Comparison of scores between the two groups using different categories for coronary artery calcification scores did not show a statistically significant difference using chi(2) analysis (P=0.088). However, there was a significant difference between dichotomized coronary artery calcification scores of lower than 100 and 100 or higher in patients with and without emphysema, respectively (P=0.013). Coexisting smoking and hypertension may contribute to the higher incidence of coronary artery calcification in emphysema patients. Symptoms of chest pain and shortness of breath were not different between the emphysema and control groups., Conclusion: Emphysema patients have a higher prevalence of significant coronary artery calcification, defined as a coronary artery calcification score higher than 100.

We investigated cortical responses and valence/arousal ratings of spider phobic, snake phobic, and healthy subjects while they were processing feared, fear-relevant, emotional neutral, and pleasant stimuli. Results revealed significantly larger amplitudes of late ERP components (P3 and late positive complex, LPC) but not of early components (N1, P2, N2) in phobics when subjects were processing feared stimuli. This fear-associated increase of amplitudes of late ERP components in phobic subjects was maximal at centro-parietal and occipital brain sites. Furthermore, phobics but not controls rated feared stimuli to be more negative and arousing than fear-relevant, emotional neutral, and pleasant stimuli. Since late ERP components and valence/arousal ratings were only significantly increased when phobic subjects but not when healthy controls were processing feared stimuli, the present data suggest that P3 and LPC amplitudes represent useful neural correlates of the emotional significance and meaning of stimuli.

The molybdenum cofactor (Moco) consists of a unique and conserved pterin derivative, usually referred to as molybdopterin (MPT), which coordinates the essential transition metal molybdenum (Mo). Moco is required for the enzymatic activities of all Mo-enzymes, with the exception of nitrogenase and is synthesized by an evolutionary old multi-step pathway that is dependent on the activities of at least six gene products. In eukaryotes, the final step of Moco biosynthesis, i.e. transfer and insertion of Mo into MPT, is catalyzed by the two-domain proteins Cnx1 in plants and gephyrin in mammals. Gephyrin is ubiquitously expressed, and was initially found in the central nervous system, where it is essential for clustering of inhibitory neuroreceptors in the postsynaptic membrane. Gephyrin and Cnx1 contain at least two functional domains (E and G) that are homologous to the Escherichia coli proteins MoeA and MogA, the atomic structures of which have been solved recently. Here, we present the crystal structures of the N-terminal human gephyrin G domain (Geph-G) and the C-terminal Arabidopsis thaliana Cnx1 G domain (Cnx1-G) at 1.7 and 2.6 A resolution, respectively. These structures are highly similar and compared to MogA reveal four major differences in their three-dimensional structures: (1) In Geph-G and Cnx1-G an additional alpha-helix is present between the first beta-strand and alpha-helix of MogA. (2) The loop between alpha 2 and beta 2 undergoes conformational changes in all three structures. (3) A beta-hairpin loop found in MogA is absent from Geph-G and Cnx1-G. (4) The C terminus of Geph-G follows a different path from that in MogA. Based on the structures of the eukaryotic proteins and their comparisons with E. coli MogA, the predicted binding site for MPT has been further refined. In addition, the characterized alternative splice variants of gephyrin are analyzed in the context of the three-dimensional structure of Geph-G., (Copyright 2001 Academic Press.)

The homo-dimeric structure of a vanadium-dependent haloperoxidase (V-BPO) from the brown alga Ascophyllum nodosum (EC 1.1.11.X) has been solved by single isomorphous replacement anomalous scattering (SIRAS) X-ray crystallography at 2.0 A resolution (PDB accession code 1QI9), using two heavy-atom datasets of a tungstate derivative measured at two different wavelengths. The protein sequence (SwissProt entry code P81701) of V-BPO was established by combining results from protein and DNA sequencing, and electron density interpretation. The enzyme has nearly an all-helical structure, with two four-helix bundles and only three small beta-sheets. The holoenzyme contains trigonal-bipyramidal coordinated vanadium atoms at its two active centres. Structural similarity to the only other structurally characterized vanadium-dependent chloroperoxidase (V-CPO) from Curvularia inaequalis exists in the vicinity of the active site and to a lesser extent in the central four-helix bundle. Despite the low sequence and structural similarity between V-BPO and V-CPO, the vanadium binding centres are highly conserved on the N-terminal side of an alpha-helix and include the proposed catalytic histidine residue (His418(V-BPO)/His404(V-CPO)). The V-BPO structure contains, in addition, a second histidine near the active site (His411(V-BPO)), which can alter the redox potential of the catalytically active VO2-O2 species by protonation/deprotonation reactions. Specific binding sites for the organic substrates, like indoles and monochlordimedone, or for halide ions are not visible in the V-BPO structure. A reaction mechanism for the enzymatic oxidation of halides is discussed, based on the present structural, spectroscopic and biochemical knowledge of vanadium-dependent haloperoxidases, explaining the observed enzymatic differences between both enzymes., (Copyright 1999 Academic Press.)

The structures of cofactor-free haloperoxidases from Streptomyces aureofaciens, Streptomyces lividans, and Pseudomonas fluorescens have been determined at resolutions between 1.9 A and 1.5 A. The structures of two enzymes complexed with benzoate or propionate identify the binding site for the organic acids which are required for the haloperoxidase activity. Based on these complexes and on the structure of an inactive variant, a reaction mechanism is proposed for the halogenation reaction with peroxoacid and hypohalous acid as reaction intermediates. Comparison of the structures suggests that a specific halide binding site is absent in the enzymes but that hydrophobic organic compounds may fit into the active site pocket for halogenation at preferential sites., (Copyright 1998 Academic Press Limited.)

The crystal structure of a lipase from the bacterium Chromobacterium viscosum ATCC 6918 (CVL) has been determined by isomorphous replacement and refined at 1.6 angstroms resolution to an R-factor of 17.8%. The lipase has the overall topology of an alpha/beta type protein, which was also found for previously determined lipase structures. The catalytic triad of the active center consists of the residues Ser87, Asp263 and His285. These residues are not exposed to the solvent, but a narrow channel connects them with the molecular surface. This conformation is very similar to the previously reported closed conformation of Pseudomonas glumae lipase (PGL), but superposition of the two lipase structures reveals several conformational differences. r.m.s. deviations greater than 2 angstroms are found for the C alpha-atoms of the polypeptide chains from His15 to Asp28, from Leu49 to Ser54 and from Lys128 to Gln158. Compared to the PGL structure in the CVL structure, three alpha-helical fragments are shorter, one beta-strand is longer and an additional antiparallel beta-sheet is found. In contrast to PGL, CVL displays an oxyanion hole, which is stabilized by the amide nitrogen atoms of Leu17 and Gln88, and a cis-peptide bond between Gln291 and Leu292. CVL contains a Ca2+, like the PGL, which is coordinated by four oxygen atoms from the protein and two water molecules.

L-2-Hydroxyisocaproate dehydrogenase (L-HicDH) from Lactobacillus confusus, a homotetramer with a molecular mass of 33 kDa per subunit, belongs to the protein family of the NAD(+)-dependent L-2-hydroxycarboxylate dehydrogenases. L-HicDH was crystallized with ammonium sulphate as precipitant in the presence of NAD+. The crystals belong to the trigonal space group P3(2)21, with a = 135.9 A and c = 205.9 A, and diffract X-rays to 2.2 A resolution. The crystal structure was solved by Patterson search and molecular replacement techniques and refined to an R-value of 21.4% (2.2 to 8 A). The final structure model contains one NAD+ molecule and one sulphate ion per subunit, with 309 water molecules. An unusual feature of this crystal structure is the deviation of the protein subunits from non-crystallographic symmetry, which is so strong that it can be detected globally by self-rotation calculations in reciprocal space. This asymmetry is especially pronounced in the environment of the active site; it is reflected also in the nicotinamide conformation of NAD+ and allows some conclusions to be drawn about the catalytic mechanism. In this context, an "inner active site loop" is identified as a structural element of fundamental functional importance. Furthermore, with knowledge of the crystal structure of L-HicDH the differences in substrate specificity between L-HicDH and the L-lactate dehydrogenases can be partly explained.

D-2-hydroxyisocaproate dehydrogenase (D-HicDH) from Lactobacillus casei is a homodimeric enzyme with a molecular mass of 74.6 kDa. It catalyzes the reduction of a wide range of 2-ketocarboxylic acids to D-2-hydroxycarboxylic acids using NADH as co-substrate. The enzyme has been crystallized by vapor diffusion using ammonium sulfate as precipitant. The crystals belong to hexagonal space group type P6(3)22 with a = b = 134.1 A, c = 124.1 A and diffract X-rays to 3.0 A resolution. Packing considerations show that there are either one or two D-HicDH monomers in the asymmetric unit.

Myocardial perfusion imaging plays an important role in the selection of patients for percutaneous transluminal coronary angioplasty (PTCA) by identifying patients with coronary artery disease and by determining the physiologic significance of a particular stenosis, and therefore, the need for PTCA. In the immediate post-PTCA period, it is useful to evaluate patients with recurrent symptoms, but frequent false-positives preclude routine application in asymptomatic patients. The most important role of myocardial perfusion imaging is the evaluation of restenosis with the first 6 months post- angioplasty. It also provides accurate information in symptomatic and asymptomatic patients and in the settings of single and multivessel PTCA and partial and complete revascularization.

The thermophile NADH oxidase from Thermus thermophilus, cloned and expressed in Escherichia coli, has been purified to homogeneity and crystallized. Three different crystal forms were found to be suitable for X-ray diffraction analysis. Crystals of the tetragonal form, grown in the presence of 25% polyethylene glycol 4000 and 0.25 M-NaCl at pH 6.6, were chosen for further analysis. These crystals belong to the space group P4(1)(3)2(1)2 with refined lattice constants of a = 94.8 A and c = 49.0 A, indicating a cell content of one monomer per asymmetric unit of the crystal. The crystals diffract to a resolution of 2.2 A.

Glucose oxidase (beta-D-glucose: oxygen 1-oxidoreductase, EC 1.1.3.4) is an FAD-dependent enzyme that catalyzes the oxidation of beta-D-glucose by molecular oxygen. The crystal structure of the partially deglycosylated enzyme from Aspergillus niger has been determined by isomorphous replacement and refined to 2.3 A resolution. The final crystallographic R-value is 18.1% for reflections between 10.0 and 2.3 A resolution. The refined model includes 580 amino acid residues, the FAD cofactor, six N-acetylglucosamine residues, three mannose residues and 152 solvent molecules. The FAD-binding domain is topologically very similar to other FAD-binding proteins. The substrate-binding domain is formed from non-continuous segments of sequence and is characterized by a deep pocket. One side of this pocket is formed by a six-stranded antiparallel beta-sheet with the flavin ring system of FAD located at the bottom of the pocket on the opposite side. Part of the entrance to the active site pocket is at the interface to the second subunit of the dimeric enzyme and is formed by a 20-residue lid, which in addition covers parts of the FAD-binding site. The carbohydrate moiety attached to Asn89 at the tip of this lid forms a link between the subunits of the dimer.

The X-ray structure determination, refinement and comparison of two crystal forms of a variant (Asn115Arg) of the alkaline protease from Bacillus alcalophilus is described. Under identical conditions crystals were obtained in the orthorhombic space group P2(1)2(1)2(1) (form I) and the rhombohedral space group R32 (form II). For both space groups the structures of the protease were solved by molecular replacement and refined at 1.85 A resolution. The final R-factors are 17.9% and 17.1% for form I and form II, respectively. The root-mean-square deviation between the two forms is 0.48 A and 0.86 A for main-chain and side-chain atoms, respectively. Due to differences in crystal lattice contacts and packing, the structures of the two crystal forms differ in intermolecular interaction affecting the local conformation of three flexible polypeptide sequences (Ser50-Glu55, Ser99-Gly102, Gly258-Ser259) at the surface of the protein. While the two overall structures are very similar, the differences are significantly larger than the errors inherent in the structure determination. As expected, the differences in the temperature factors in form I and II are correlated with the solvent accessibility of the corresponding amino acid residues. In form II, two symmetry-related substrate binding sites face each other, forming a tight intermolecular interaction. Some residues contributing to this intermolecular interaction are also found to be involved in the formation of the complex between subtilisin Carlsberg and the proteinaceous inhibitor eglin C. This demonstrates that the two symmetry-related molecules interact with each other at the same molecular surface area that is used for binding of substrates and inhibitors.

A modified version of the human pancreatic trypsin inhibitor (PSTI), generated in a protein-design project, has been crystallized in spacegroup P4(3) with lattice constants a = 40.15 A, c = 33.91 A. The structure has been solved by molecular replacement. Refinement of the structure by simulated annealing and conventional restrained least-squares yielded for 8.0 to 2.3 A data a final R-value of 19.1%. Differences to the known structures of porcine PSTI complexed with trypsinogen and modified human PSTI complexed with chymotrypsinogen occur at the flexible N-terminal part of the molecule. These differences are influenced by crystal packing, as are low temperature factors for the binding loop. The geometry of the binding loop is similar to the complexed structures.

The dimeric glucose oxidase from Penicillium amagasakiense was deglycosylated, purified and crystallized as a complex with its coenzyme FAD. Deglycosylation and purification to isoelectric homogeneity were shown to be an important prerequisite step to obtain crystals suitable for X-ray investigations. Crystals of the deglycosylated enzyme were reproducibly grown using ammonium sulfate as precipitant at pH 7.4 to 7.5. Crystals diffract to at least 2.0 A resolution and belong to the orthorhombic space group P2(1)2(1)2(1), with refined lattice constants of a = 59.3 A, b = 136.3 A and c = 156.7 A. Assuming two monomers (approximately 135 kDa) per asymmetric unit the Vm value is 2.3 A3/Da.

Variants of the human pancreatic secretory trypsin inhibitor (PSTI) have been created during a protein design project to generate a high-affinity inhibitor with respect to some serine proteases other than trypsin. Two modified versions of human PSTI with high affinity for chymotrypsin were crystallized as a complex with chymotrypsinogen. Both crystallize isomorphously in space group P4(1)2(1)2 with lattice constants a = 84.4 A, c = 86.7 A and diffract to 2.3 A resolution. The structure was solved by molecular replacement. The final R-value after refinement with 8.0 to 2.3 A resolution data was 19.5% for both complexes after inclusion of about 50 bound water molecules. The overall three-dimensional structure of PSTI is similar to the structure of porcine PSTI in the trypsinogen complex (1TGS). Small differences in the relative orientation of the binding loop and the core of the inhibitors indicate flexible adaptation to the proteases. The chymotrypsinogen part of the complex is similar to chymotrypsin. After refolding induced by binding of the inhibitor the root-mean-square difference of the active site residues A186 to A195 and A217 to A222 compared to chymotrypsin was 0.26 A.

Deglycosylation was shown to be an important prerequisite step for the crystallization of glucose oxidase from Aspergillus niger. Whereas the glycosylated enzyme could not be crystallized, crystals of the deglycosylated enzyme suitable for X-ray diffraction analysis were reproducibly grown in the presence of 1.6 M-ammonium sulphate and octanetriol at pH 5.3 to 5.6. The crystals belong to the space group P3(1)21 or P3(2)21 with refined lattice constants of a = 66.5 A and c = 214.4 A, indicating a cell content of one monomer per asymmetric unit of the crystal. Crystals diffract to at least 2.5 A resolution. Cleavage of 95% of its carbohydrate moiety affected the kinetics of glucose oxidation, stability at low pH and some electrophoretic properties of glucose oxidase, such as molecular mass and the number of isoelectric forms. However, other properties, such as thermal stability, pH and temperature optima of activity were not affected.

A new cubic crystal form (a = 445.1 A) of space group P23 is reported for human rhinovirus R14. There are four particles per unit cell, each situated on a crystallographic 3-fold axis. The orientation of these particles has been determined with a rotation function and their approximate positions have been derived from a Patterson map. The crystals diffract to at least 2.8 A resolution. Limitations to the possible surface features of the virus are set by a comparison of the cubic and orthorhombic crystal forms.