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Ubiquitination is a crucial post-translational modification; however, the functions of ubiquitin-coding genes remain unclear. UBA52 encodes a fusion protein comprising ubiquitin at the N-terminus and ribosomal protein L40 (RPL40) at the C-terminus. Here we showed that Uba52-deficient mice die during embryogenesis. UBA52-deficient cells exhibited normal levels of total ubiquitin. However, UBA52-deficient cells displayed decreased protein synthesis and cell-cycle arrest. The overexpression of UBA52 ameliorated the cell-cycle arrest caused by UBA52 deficiency. Surprisingly, RPL40 expression itself is insufficient to regulate cyclin D expression. The cleavage of RPL40 from UBA52 was required for maintaining protein synthesis. Furthermore, we found that RPL40 formed a ribosomal complex with ubiquitin cleaved from UBA52. UBA52 supplies RPL40 and ubiquitin simultaneously to the ribosome. Our study demonstrated that the ubiquitin-coding gene UBA52 is not just an ubiquitin supplier to the ubiquitin pool but is also a regulator of the ribosomal protein complex. These findings provide novel insights into the regulation of ubiquitin-dependent translation and embryonic development.

Prior reports have revealed that complete revascularization (CR) by percutaneous coronary intervention (PCI) decreased ischemic events. However, little is known about the efficacy of CR using PCI in elderly patients with multi-vessel coronary artery disease (CAD). We evaluated the 1-year effectiveness of CR-PCI in elderly patients (≥75years old) with multi-vessel CAD.The SHINANO Registry, a prospective, observational, multi-center, all-comer cohort study, has enrolled 1923 patients. From this registry, we recruited 322 elderly patients with multi-vessel CAD. The primary endpoint was major adverse cardiovascular events ([MACE]: all-cause mortality, myocardial infarction, and stroke).Of the 322 elderly patients with multi-vessel CAD, 165 (51.2%) received CR and 157 (48.8%) received incomplete revascularization (ICR). MACE occurred in 44 (13.7%) patients. The incidence of MACE by survival analysis was significantly lower in the CR group than in the ICR group (7.4% vs. 21.1%, p0.001). On multivariable Cox proportional hazards analysis of age, sex, and acute coronary syndrome (ACS), ACS and CR were independent predictors of MACE (hazard ratio [HR], 2.49; 95% confidence interval [CI], 1.29-4.80; p=0.007, HR, 0.40; 95% CI, 0.20-0.77; p=0.007, respectively). In propensity score matching of age, sex, previous heart failure, previous intracranial bleeding, ACS, and body mass index, the MACE rate was significantly lower in the CR group than in the ICR group (7.2% vs. 18.4%, p=0.015).Even in elderly patients over 75years old with multi-vessel CAD, CR-PCI appears to suppress mid-term ischemic events.

High morbidity and mortality associated with human cases of highly pathogenic avian influenza (HPAI) viruses, including H5N1 influenza virus, have been reported. The purpose of the present study was to evaluate the antiviral effects of peramivir against HPAI viruses. In neuraminidase (NA) inhibition and virus replication inhibition assays, peramivir showed strong inhibitory activity against H5N1, H7N1 and H7N7 HPAI viruses with sub-nanomolar activity in enzyme assays. In H5N1 viruses containing the NA H275Y mutation, the antiviral activity of peramivir against the variant was lower than that against the wild-type. Evaluation of the in vivo antiviral activity showed that a single intravenous treatment of peramivir (10 mg/kg) prevented lethality in mice infected with wild-type H5N1 virus and also following infection with H5N1 virus with the H275Y mutation after a 5 day administration of peramivir (30 mg/kg). Furthermore, mice injected with peramivir showed low viral titers and low levels of proinflammatory cytokines in the lungs. These results suggest that peramivir has therapeutic activity against HPAI viruses even if the virus harbors the NA H275Y mutation.

Fused deposition modeling (FDM) 3–Dimensional (3D) printing is becoming an increasingly important technology in the pharmaceutical sciences, since it allows the manufacture of personalized oral dosage forms by deposition of thin layers of material. Here, a filament extruder was used to obtain filaments of polyvinyl alcohol (PVA) containing paracetamol or caffeine appropriate for 3D printing. The filaments were used to manufacture caplets for oral administration by FDM 3D printing, with the aim of evaluating the effect of the internal structure (micropore volume), drug loading and composition on drug dissolution behaviour. Micropore volume of the caplets was primarily determined by the presence of large pores due to gaps in the printed layers/net while printing, and the porosity of the caplets was 10 fold higher than the porosity of the extruded filament. Dynamic dissolution drug release tests on the caplets in biorelevant bicarbonate media revealed distinctive release profiles, which were dependent on drug solubility and drug loading. Porosity of the caplets did not help to predict the different drug release profiles. This study confirms the potential of 3D printing to fabricate caplets and helps to elucidate which factors influence drug release from this type of new dosage form.

Canine embryos (8-cell to blastocyst stages) frozen-thawed using the slow-freezing method with glycerol (four recipients) or dimethyl sulfoxide (three recipients) as a cryoprotectant and vitrified-warmed using the Cryotop method (five recipients) were surgically transferred into the unilateral uterine horn of recipient bitches. As a result, the morphology of embryos frozen-thawed using the slow-freezing method was judged to be normal, but no conception occurred in any of the recipient bitches. Two of the five bitches that received transferred embryos (morula to early blastocyst stages) vitrified-warmed using the Cryotop method became pregnant and produced normal pups (1/9 embryos, 11.1% and 1/6 embryos, 17.0%). It was concluded that the Cryotop method was more appropriate for canine embryo cryopreservation than the slow-freezing method, which is used for the cryopreservation of embryos of other mammalian species.

We collected semen from a male Amur leopard cat using the transrectal electroejaculation method and investigated the semen qualities for about four years. In addition, the influence of the season on the spermatogenic function of the Amur leopard cat was investigated with regard to the semen qualities, testicular volume and serum testosterone level. As a result, we could collect semen with good sperm qualities that would be useable for artificial insemination. Some seasonality was noted in the testicular volume and serum testosterone level. We clarified that the semen qualities were favorable before and during the female breeding season compared with those after the breeding season.

Kaguya gamma-ray spectrometer measured thorium (Th) distribution on the lunar farside with the highest sensitivity among past gamma-ray remote sensing missions. The newly obtained Th map has revealed that two regions near the equator on the farside have the lowest Th abundances. We found that the variation of the Th abundance perfectly correlates with the crustal thickness in the farside and the southern nearside, and it could be a result of the crystallization of the lunar magma ocean.

This is a short note on the relation between the graded stable derived categories of 14 exceptional unimodal singularities and the derived category of K3 surfaces obtained as compactifications of the Milnor fibers. As a corollary, we obtain a basis of the numerical Grothendieck group similar to the one given by Ebeling and Ploog (arXiv:0809.2738)., Comment: 21 pages, 4 figures; v2: corrected an error pointed out by the referee

Drug-induced liver injury (DILI) is a major safety concern in drug development and clinical practice. The pathogenesis of DILI usually involves the participation of the parent drug or metabolites that either affect cellular function or elicit an immune response. However, the mechanisms leading to DILI are unknown in most cases. Methimazole (MTZ) is used as an antithyroid drug and is well known to have induced liver injuries such as cholestatic hepatitis in a small number of human cases. Immune-mediated reactions were also suggested to play a role in MTZ-induced acute liver injury, but the mechanism underlying this process has not been elucidated. To address this issue, we measured plasma aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, hepatic glutathione levels, hepatic expression of CD4⁺ Th cell-related transcriptional factors, cytokines and chemokines, plasma interleukin (IL)-4 levels and histopathological changes in the liver following MTZ (450 mg kg⁻¹ , p.o.) administration in mice. The hepatic expression of mRNA for Th2 cell-related factors, such as GATA-binding protein, macrophage inflammatory protein-2 (MIP-2) and plasma IL-4 levels, as well as plasma AST and ALT levels, was significantly increased in mice treated with MTZ. These changes were markedly enhanced by pre-treatment with L-buthionine sulfoximine (3 mmol kg⁻¹, i.p.) and MTZ (15 mg kg⁻¹, p.o.). Neutralization of IL-4 using a monoclonal anti-mouse IL-4 antibody (100 µg/mouse, single i.p.) suppressed the hepatotoxic effect of MTZ. In conclusion, this report is the first to demonstrate that Th2 cytokine-mediated immune responses are involved in MTZ-induced acute liver injury in mice.

A Beagle dog (3 years old) that ejaculated high percentages (mean ± SE: 29.1 ± 1.2%) of sperm with a knobbed acrosome abnormality and a low number of sperm and that also had a low plasma testosterone (T) level was given 10 subcutaneous injections of 1 µg of gonadotropin-releasing hormone analogue (GnRH-A) at 3-day intervals. The plasma T level and number of sperm increased 12-14 weeks after the first injection. Although the percentages of sperm with knobbed acrosome abnormality did not change after the GnRH-A therapy, the number of sperm and percentage of actively motile sperm increased after the therapy, and a bitch gave birth to 5 healthy puppies after intravaginal artificial insemination with fresh semen collected 14 weeks after the first injection.

著者人数: 14名, Accepted: 2012-01-04, 資料番号: SA1004214000

The responses of a piezoelectric lead zirconate titanate (PZT) element to hypervelocity collisions were experimentally studied. In this study, the particles of masses ranging from 0.3 to 10 fg were made to collide with PZT at velocities between 20 and 96 km/s. The amplitude and the corresponding rise time of the single-pulse output signals that were produced in the piezoelectric PZT element were measured to determine the possible collision states. The results revealed an apparently multimodal output; three classes were assumed to be involved in the pulse formation mechanism. The amplitude and rise time were sensitive to the collision velocity. The multimodal behavior implied that the PZT-based cosmic dust detectors should be calibrated according to the class they belong to.

Drug-induced liver injury is a growing concern for pharmaceutical companies and patients because numerous drugs have been linked to hepatotoxicity and it is the most common reason for a drug to be withdrawn. Flutamide rarely causes liver dysfunction in humans, and immune allergic reactions have been suggested in some cases. In this study, we investigated the mechanisms of flutamide-induced liver injury in BALB/c mice. Plasma alanine aminotransferase and aspartate aminotransferase levels were significantly increased 3, 6 and 9 h after flutamide (1500 mg kg⁻¹ , p.o.) administration. The biomarker for oxidative stress was not changed, but Th2-dominant immune-related factors, such as interleukin (IL)-4, IL-5, STAT6 and GATA-binding protein (GATA)-3, were induced in flutamide-administered mice. The pre-administration of monoclonal-IL-4 antibody suppressed the hepatotoxicity of flutamide. In addition, we investigated the effect of 13,14-dihydro-15-keto-PGD₂ (DK-PGD₂; 10 µg per mouse, i.p.) administration on flutamide-induced acute liver injury. Coadministration of DK-PGD₂ and flutamide resulted in a significant increase in alanine aminotransferase and a remarkable increase of macrophage inflammatory protein-2. In conclusion, we demonstrated that flutamide-induced acute liver injury is mediated by Th2-dominant immune responses in mice.

In present study, the effect of the ultrasound wave stimulation in acceleration of the osteocondutibity of bio-active pure titanium was investigated. Pure titanium (99.9% Ti) processed in chemical and heat treatments was used as a specimen, and soaked in simulated body fluid (SBF) under pulsed ultrasound wave for the planned time periods. The surface of samples was observed using scanning electron microscopy (SEM), X-ray diffraction, etc, to assess the state of hydroxyapatite formation. SEM images showed that a richer and finer layer of calcium and phosphate compounds covered the titanium surface in the ultrasound wave group as compared with the non-ultrasound group. The measurements of mass of specimens also indicated the efficiency of ultrasound waves for hydroxyapatite formation. These findings suggest that the nucleation and crystallization of apatite on bio-active material surfaces might be promoted by micro-moving and cavitation of low-intensity pulsed ultrasound waves. We propose that pulsed ultrasound stimulation has a great potential for further improvement of osteointegration and osteoconductivity for medical bio-active implants.

The effect intra-articular administration of the hyaluronic acid (HA) for the treatment of osteoarthritis knee joint has been widely recognized. However, the efficacy of mechanical tribology function of HA as a viscosupplementation to joint lubrication has not yet been confirmed. To examine in vivo the effects of enhancement of tribology function of joint lubricant for the treatment of osteoarthritis, animal experiment was performed. Experiment osteoarthritis of the both of knee joints of three rabbits was induced by meniscsectomy and cartilage defects. These animals were subjected to monthly intra-articular cartilage injection of high viscous polyethylene glycol (PEG) to one side knee joint (another side was not injected as a control knee), and were sacrificed at 3 month after operation. The extracted knee joints were evaluated macroscopically and histologically. As a results, the PEG injected knees had a tendency to demonstrate less damage of articular cartilage compared with the control group. This finding indicated that the development and addition of mechanical lubricant for the preventing of articular cartilage wear could have potential to be one of effective treatment for arthritis.

本研究はThink/No-Thinkパラダイム(Anderson & Green，2001)を用いて，ポジティブ画像記憶の意図的抑制が可能かを検討した．方法は学習段階，Think/No-Think段階，最終テスト段階の3段階から構成されていた．学習段階において，実験参加者(N=34)は手がかり(ニュートラル単語)とターゲット(ポジティブもしくはニュートラル画像)の刺激ペアを学習した．その後，正解率が80%を超えるまで手がかり再認テストを行った．Think/No-Think段階では，手がかりのみが反復して提示された(0回，6回，12回)．0回反復のペアは記憶の促進と記憶の抑制を評価するためのベースラインとして使用された．赤色の手がかりに対して，実験参加者はペアとなっていたターゲットを思い出した．緑色の手がかりに対しては，ペアとなっていたターゲットを抑制した．最終テスト段階では，手がかり再生テストを行った．結果，ポジティブ，ニュートラル画像共に，抑制されたターゲットはベースラインのターゲットよりも記憶成績が低かった．画像記憶の抑制が頑健であること，そして感情価によって抑制中の主観的体験が異なることが示唆された．

Hippocampal atrophy, which is a component of hippocampal sclerosis and recognized commonly in human intractable epilepsy, is controversial in canine epilepsy. We examined the hippocampal volume in 58 epileptic dogs and 35 control dogs using magnetic resonance (MR) images, and calculated the relative hippocampal volume asymmetry of the right and left hippocampus. Subjectively, there were visible MR imaging abnormalities in seven of the 58 epileptic dogs (12%). The hippocampel volume asymmetric ratio of epileptic dogs (5.84 +/- 4.47%) was significantly greater than that of control dogs (1.62 +/- 0.88%). Using a cutoff threshold asymmetric ratio of 6% that is indicated in human epilepsy, 28 epileptic dogs (48%) were characterized as having unilateral hippocampus atrophy. The hippocampal volume asymmetry ratio cannot be used to detect bilateral atrophy. In conclusion, although less frequent than that observed in human epilepsy patients, hippocampal atrophy may occur in canine epilepsy.

The Kaguya gamma-ray spectrometer (KGRS) has great potential to precisely determine the absolute abundances of natural radioactive elements K, Th and U on the lunar surface because of its excellent spectroscopic performance. In order to achieve the best performance of the KGRS, it is important to know the spatial response function (SRF) that describes the directional sensitivity of the KGRS. The SRF is derived by a series of Monte Carlo simulations of gamma-ray transport in the sensor of the KGRS using the full-fledged simulation model of the KGRS, and is studied in detail. In this paper, the method for deriving absolute abundance of natural radioactive elements based on the SRF is described for the analysis of KGRS data, which is also applicable to any gamma-ray remote sensings. In the preliminary analysis of KGRS data, we determined the absolute abundances of K and Th on the lunar surface without using any previous knowledge of chemical information gained from Apollo samples, lunar meteorites and/or previous lunar remote sensings. The results are compared with the previous measurements and the difference and the correspondence are discussed. Future detailed analysis of KGRS data will provide new and more precise maps of K, Th and U on the lunar surface.

Due to its excellent biocompatibility and mechanical properties, various different applications of polyvinyl alcohol-hydrogels (PVA-H) has been attempted in many fields. In the field of orthopedic surgery, we have been engaged for long time in research on the clinical applications of PVA-H as a artificial cartilage, and have performed many basic experiments on the mechanical properties, synthesis of PVA-H, and developed orthopedic implants using PVA-H. From these studies, many applications of artificial articular cartilage, intervertbral disc and artificial meniscus etc. have been developed. This review will present the overview of the applications and recent advances of PVA-H cartilages, and discuss clinical potential of PVA-H for orthopedics implant.

Duchenne muscular dystrophy (DMD) is a devastatingmuscle disorder that is characterized by progressive muscle necrosis,fibrosis, and fatty infiltration. To examine the temporospatial pathologicalchanges, a noninvasive evaluation method such as magnetic resonanceimaging (MRI) is needed. The aim of this study was to precisely assessmuscle necrosis and inflammation based on a sequence of T2-weightedimaging (T2WI), gadolinium-enhanced imaging, and selective fat suppres-sion, chemical shift selective T2-weighted imaging (CHESS-T2WI), on a 3.0-Tesla MRI unit in 3-month-old and 7-year-old dogs with canine X-linkedmuscular dystrophy (CXMD J ), a suitable animal model for DMD. The resultsshow that CHESS-T2WI was more sensitive and useful from the early to latestages of CXMD J than T2WI or contrast enhancement imaging in theevaluation of muscle necrosis, because these latter sequences can beinfluenced by fatty infiltration or interstitial connective tissues. Muscle Nerve 40: 815–826, 2009

When a Fourier series is used to approximate a periodic and piecewise smooth function with a jump discontinuity, an overshoot at the discontinuity occurs and is called Gibbs phenomenon. For explaining understandably and systematically this Gibbs phenomenon from the educational point of view , the representation method is proposed using an accordion-like folding convergence of the extremum values for the partial sums of its Fourier series at the discontinuity. Using an integration by parts to obtain Fourier coefficients and rearranging its Fourier series, the expansions are devised to show explicitly the overshoots at the discontinuities for any functions with jump discontinuities.

Resistance passage studies were conducted with five INIs (integrase inhibitors) that have been tested in clinical trials to date: a new naphthyridinone-type INI S/GSK-364735, raltegravir, elvitegravir, L-870,810 and S-1360. In establishing the passage system and starting from concentrations several fold above the EC50 value, resistance mutations against S-1360 and related diketoacid-type compounds could be isolated from infected MT-2 cell cultures from day 14 to 28. Q148R and F121Y were the two main pathways of resistance to S/GSK-364735. Q148R/K and N155H, which were found in patients failing raltegravir treatment in Phase IIb studies, were observed during passage with raltegravir with this method. The fold resistance of 40 mutant molecular clones versus wild type virus was compared with these five INIs. The overall resistance pattern of S/GSK-364735 was similar to that of raltegravir and other INIs. However, different fold resistances of particular mutations were noted among different INIs, reflecting a potential to develop INIs with distinctly different resistant profiles.

The high precision gamma-ray spectrometer (GRS) is scheduled to be launched on the lunar polar orbiter of the SELENE mission in 2007. The GRS consists of a large Ge crystal as a main detector and massive bismuth germanate crystals as an anticoincidence detector. A Stirling cryocooler was adopted in cooling the Ge detector. The flight model of SELENE GRS has been completed and an energy resolution of 3.0 keV (FWHM) at 1.332 MeV has been achieved. The spectrometer aims to observe nuclear line gamma rays emitted from the lunar surface in a wide energy range from 100 keV to 12 MeV for one year and more to obtain chemical composition on the entire lunar surface. The gamma-ray data enable us to study lunar geoscience problems including crust and mantle composition, and volatile reservoirs at polar regions.

Assuming different abundances of the Apollo lunar soil samples and the average spectrum of galactic cosmic ray protons, energy spectra of neutrons and gamma rays and emission rates of gamma-ray lines from major elements have been estimated by using the reviewed Monte Carlo simulation library Geant4 and nuclear data. Previously, such libraries were not able to reproduce gamma-ray lines properly for the planetary application. Results clearly show that the emission rate of gamma rays heavily depends not only on the chemical abundance but also neutron flux within the lunar subsurface. While the intensities of gamma-ray lines are mostly proportional to elemental abundances, the intensity per unit elemental abundance can vary. Such a complex correlation is attributed to the change in neutron flux within the lunar subsurface and petrological restriction of elemental variation.

The naphthyridinone GSK364735 potently inhibited recombinant human immunodeficiency virus type 1 (HIV-1) integrase in a strand transfer assay (mean 50% inhibitory concentration ± standard deviation, 8 ± 2 nM). As expected based on the structure of the drug, it bound competitively with another two-metal binding inhibitor ( K d [binding constant], 6 ± 4 nM). In a number of different cellular assays, GSK364735 inhibited HIV replication with potency at nanomolar concentrations (e.g., in peripheral blood mononuclear cells and MT-4 cells, 50% effective concentrations were 1.2 ± 0.4 and 5 ± 1 nM, respectively), with selectivity indexes of antiviral activity versus in-assay cytotoxicity of at least 2,200. When human serum was added, the antiviral potency decreased (e.g., a 35-fold decrease in the presence of 100% human serum was calculated by extrapolation from the results of the MT-4 cell assay). In cellular assays, GSK364735 blocked viral DNA integration, with a concomitant increase in two-long-terminal-repeat circles. As expected, this integrase inhibitor was equally active against wild-type viruses and mutant viruses resistant to approved drugs targeting either reverse transcriptase or protease. In contrast, some but not all viruses resistant to other integrase inhibitors were resistant to GSK364735. When virus was passaged in the presence of the inhibitor, we identified resistance mutations within the integrase active site that were the same as or similar to mutations arising in response to other two-metal binding inhibitors. Finally, either additive or synergistic effects were observed when GSK364735 was tested in combination with approved antiretrovirals (i.e., no antagonistic effects were seen). Thus, based on all the data, GSK364735 exerted potent antiviral activity through the inhibition of viral DNA integration by interacting at the two-metal binding site within the catalytic center of HIV integrase.

Experimental study of soot formation in DME jet diffusion flame was performed to investigate the influence of the oxygen contained in the molecular structure of DME. The amount of soot in the flame was measured using the transmitted light attenuation method, and was compared with that of the partial premixing methane flame under the high temperature and various surrounding oxygen conditions. The results showed that the amount of soot in the DME diffusion flame is much less than that of methane flame in most conditions, and the partially premixied methane flame of φ=7 that is substantially equal in equivalent ratio with DME diffusion flame generated 3-10 times as much soot as DME. This result suggests that the suppression of soot formation in DME flame is not due to the decrease of substantial equivalent ratio with contained oxygen, but attributed to the molecular structure of DME. The volume of the methane that is generated by primary pyrolysis of DME might have considerable influence on the generated soot amount.

症例は72歳，女性．腸閉塞の診断にて当院入院となった．精密検査にてS状結腸癌による腸閉塞と診断し，S状結腸切除術を施行した．開腹所見にて腹腔内に径2-3mmの腹膜小結節を無数に認め，肉眼的に腹膜播種と診断した．術後の病理検査結果にて腹膜結節は中心性乾酪壊死と周囲にLanghans型巨細胞を伴う肉芽腫であり，S状結腸癌に合併した結核性腹膜炎と診断した．術後抗結核剤による治療を開始した．結核性腹膜炎は稀な疾患であるが，悪性腫瘍と合併した場合に癌性腹膜炎と鑑別できない危険性があり，両疾患に対する適切な治療機会を逸する可能性もある．粟粒大の腹膜結節を認めた場合には念頭に置くべき疾患と考えられる．

Objective Smoking is associated with gastroesophageal reflux disease (GERD). Varenicline, a nicotinic receptor partial agonist, is used to aid smoking cessation. The purpose of this study was to prospectively examine the long-term benefits of smoking cessation on GERD and health-related quality of life (HR-QOL). Methods Patients treated with varenicline were asked to fill out a self-report questionnaire about their smoking habits, gastrointestinal symptoms, and HR-QOL before and 1 year after smoking cessation. The prevalence of GERD, frequency of symptoms, and HR-QOL scores were compared. We also investigated associations between clinical factors and newly-developed GERD. Results A total of 141 patients achieved smoking cessation (success group) and 50 did not (failure group) at 1 year after the treatment. The GERD improvement in the success group (43.9%) was significantly higher than that in the failure group (18.2%). The frequency of reflux symptoms significantly decreased only in the success group. There were no significant associations between newly developed GERD and clinical factors including increased body mass index and successful smoking cessation. HR-QOL significantly improved only in the success group. Conclusions Smoking cessation improved both GERD and HR-QOL. Smoking cessation should be recommended for GERD patients.

Disulfide bonds play an important role in protein stability and function. Here, we describe a general procedure for generating disulfide-linked dimers and multimers of proteins of known crystal structures. An algorithm was developed to predict sites in a protein compatible with intermolecular disulfide formation with neighboring molecules in the crystal lattice. A database analysis was carried out on 46 PDB coordinates to verify the general applicability of this algorithm to predict intermolecular disulfide linkages. On the basis of the predictions from this algorithm, mutants were constructed and characterized for a model protein, thioredoxin. Of the five mutants, as predicted, in solution four formed disulfide-linked dimers while one formed polymers. Thermal and chemical denaturation studies on these mutant thioredoxins showed that three of the four dimeric mutants had similar stability to wild-type thioredoxin while one had lower stability. Three of the mutant dimers crystallized readily (in four to seven days) in contrast to the wild-type protein, which is particularly difficult to crystallize and takes more than a month to form diffraction-quality crystals. In two of the three cases, the structure of the dimer was exactly as predicted by the algorithm, while in the third case the relative orientation of the monomers in the dimer was different from the predicted one. This methodology can be used to enhance protein crystallizability, modulate the oligomerization state and to produce linear chains or ordered three-dimensional protein arrays.