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Degradation models based on heterogeneous Wiener processes are commonly used to assess information on the lifetime of highly reliable products. An optimal test plan given limited resources is generally obtained using numerical methods for heterogeneous Wiener processes. However, numerical searches for optimal test plans have the disadvantage of being time‐consuming and may provide unclear explanations for the findings. To overcome these difficulties, we derive an explicit expression for decision variables (such as the termination time, number of measurements, and sample size) of D$$ D $$‐ and V$$ V $$‐optimal test plans with cost constraints. The theoretical results not only ensure that the optimal test plan is found, but also provide clear insights into the decision variables affected by model parameters and experimental costs. Some numerical examples are presented to support the efficiency and applicability of the optimal test plans. [ABSTRACT FROM AUTHOR]

Degradation tests are widely used to assess the lifetime information of highly reliable products. Before conducting a degradation test, the fundamental issues in regard to decision variables are to determine how many sample sizes are needed, how long samples need to be tested, and how many measurements are taken, particularly in early experiments with limited budgets. By minimizing the approximate variance of the estimated qth quantile of the product's lifetime distribution with a cost constraint, profile optimum planning (POP) is proposed to provide a systematic solution to these decision variables under mild conditions. Based on the derived theoretical results, the use of flow charts efficiently simplifies complex optimization problems in practical applications. In addition, sensitivity analysis is studied to elucidate the effect of how the uncertainty in POP can be divided and allocated to the experimental costs and model parameters. Technical derivations, flow charts, and analysis results are available online as Supporting Information. [ABSTRACT FROM AUTHOR]

Stochastic processes are widely used to analyze degradation data, and the Gaussian process is a particularly common one. In this article, we propose a robust statistical model using a Student-t process to assess the lifetime information of highly reliable products. This model is statistically plausible and demonstrates a substantially improved fit when applied to real data. A computationally accurate approach is proposed to calculate the first-passage-time density function of the Student-t degradation-based process; related properties are investigated as well. In addition, this article provides parameter estimation using the EM-type algorithm and a simple model-checking procedure to evaluate the appropriateness of the model assumptions. Several case studies are performed to demonstrate the flexibility and applicability of the proposed model with random effects and explanatory variables. Technical details, datasets, and R codes are available as . [ABSTRACT FROM AUTHOR]

Abstract: We report the first X‐ray crystallographic structure of the “head‐to‐middle” prenyltransferase, isosesquilavandulyl diphosphate synthase, involved in biosynthesis of the merochlorin class of antibiotics. The protein adopts the ζ or cis‐prenyl transferase fold but remarkably, unlike tuberculosinol adenosine synthase and other cis‐prenyl transferases (e.g. cis‐farnesyl, decaprenyl, undecaprenyl diphosphate synthases), the large, hydrophobic side chain does not occupy a central hydrophobic tunnel. Instead, it occupies a surface pocket oriented at 90° to the hydrophobic tunnel. Product chain‐length control is achieved by squeezing out the ligand from the conventional allylic S1 binding site, with proton abstraction being achieved using a diphosphate‐Asn‐Ser relay. The structures revise and unify our thinking as to the mechanism of action of many other prenyl transferases and may also be of use in engineering new merochlorin‐class antibiotics. [ABSTRACT FROM AUTHOR]

We report the first X-ray structure of the unique 'head-to-middle' monoterpene synthase, lavandulyl diphosphate synthase (LPPS). LPPS catalyzes the condensation of two molecules of dimethylallyl diphosphate (DMAPP) to form lavandulyl diphosphate, a precursor to the fragrance lavandulol. The structure is similar to that of the bacterial cis-prenyl synthase, undecaprenyl diphosphate synthase (UPPS), and contains an allylic site (S1) in which DMAPP ionizes and a second site (S2) which houses the DMAPP nucleophile. Both S-thiolo-dimethylallyl diphosphate and S-thiolo-isopentenyl diphosphate bind intact to S2, but are cleaved to (thio)diphosphate, in S1. His78 (Asn in UPPS) is essential for catalysis and is proposed to facilitate diphosphate release in S1, while the P1 phosphate in S2 abstracts a proton from the lavandulyl carbocation to form the LPP product. The results are of interest since they provide the first structure and structure-based mechanism of this unusual prenyl synthase. [ABSTRACT FROM AUTHOR]

ABSTRACT Xylanases are capable of decomposing xylans, the major components in plant cell wall, and releasing the constituent sugars for further applications. Because xylanase is widely used in various manufacturing processes, high specific activity, and thermostability are desirable. Here, the wild-type and mutant (E146A and E251A) catalytic domain of xylanase from Thermoanaerobacterium saccharolyticum JW/SL-YS485 (TsXylA) were expressed in Escherichia coli and purified subsequently. The recombinant protein showed optimal temperature and pH of 75°C and 6.5, respectively, and it remained fully active even after heat treatment at 75°C for 1 h. Furthermore, the crystal structures of apo-form wild-type TsXylA and the xylobiose-, xylotriose-, and xylotetraose-bound E146A and E251A mutants were solved by X-ray diffraction to high resolution (1.32-1.66 Å). The protein forms a classic (β/α)8 folding of typical GH10 xylanases. The ligands in substrate-binding groove as well as the interactions between sugars and active-site residues were clearly elucidated by analyzing the complex structures. According to the structural analyses, TsXylA utilizes a double displacement catalytic machinery to carry out the enzymatic reactions. In conclusion, TsXylA is effective under industrially favored conditions, and our findings provide fundamental knowledge which may contribute to further enhancement of the enzyme performance through molecular engineering. Proteins 2013; 81:1256-1265. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The endoglucanase EglA from Piromyces rhizinflata found in cattle stomach belongs to the GH5 family of glycoside hydrolases. The crystal structure of the catalytic domain of EglA shows the (β/α)8-barrel fold typical of GH5 enzymes. Adjacent to the active site of EglA, a loop containing a disulfide bond not found in other similar structures may participate in substrate binding. Because the active site was blocked by the N-terminal His tag of a neighbouring protein molecule in the crystal, enzyme-substrate complexes could not be obtained by soaking but were prepared by cocrystallization. The E154A mutant structure with a cellotriose bound to the −3, −2 and −1 subsites shows an extensive hydrogen-bonding network between the enzyme and the substrate, along with a stacking interaction between Trp44 and the −3 sugar. A possible dimer was observed in the crystal structure, but retention of activity in the E242A mutant suggested that the enzyme probably does not function as a dimer in solution. On the other hand, the first 100 amino acids encoded by the original cDNA fragment are very similar to those in the last third of the (β/α)8-barrel fold, indicating that EglA comprises at least two catalytic domains acting in tandem. [ABSTRACT FROM AUTHOR]

The article presents a diagram related to the article "Structure and Function of a "Head-to-Middle" Prenyltransferase: Lavandulyl Diphosphate Synthase" by Y. Ma and colleagues in the current issue.

Die irreguläre Terpensynthase Lavandulyldiphosphatsynthase (LPPS) produziert die Vorstufe des kommerziell wichtigen Lavendelduftstoffs Lavandulol durch einen ungewöhnlichen “Kopf ‐ zur ‐ Mitte” ‐ Mechanismus. E. Oldfield, R. ‐ T. Guo, Y. Ma et al. präsentieren in der Zuschrift auf S. 4799 ff. die erste Struktur von LPPS im Komplex mit seinem Substrat. Die schmetterlingsförmige Struktur ähnelt der von Proteinen, die an der Biosynthese bakterieller Zellwände beteiligt sind, und führt zu einem einzigartigen Kondensationsmechanismus, der von allgemeinem Interesse im Kontext der Terpensynthese ist. [ABSTRACT FROM AUTHOR]

Cellulose is the major component of the plant cell wall and the most abundant renewable biomass on earth, and its decomposition has proven to be very useful in many commercial applications. Endo-1,4-β- d -glucanase (EC 3.2.1.4; endoglucanase), which catalyzes the random hydrolysis of 1,4-β-glycosidic bonds of the cellulose main chain to cleave cellulose into smaller fragments, is the key cellulolytic enzyme. An endoglucanase isolated from Aspergillus aculeatus F-50 (FI-CMCase), which is classified into the glycoside hydrolase (GH) family 12, was demonstrated to be effectively expressed in the industrial strain Pichia pastoris . Here, the crystal structure and complex structures of P. pastoris -expressed FI-CMCase were solved to high resolution. The overall structure is analyzed and compared to other GH12 members. In addition, the substrate-surrounding residues were engineered to search for variants with improved enzymatic activity. Among 14 mutants constructed, one with two-fold increase in protein expression was identified, which possesses a potential to be further developed as a commercial enzyme product. [ABSTRACT FROM AUTHOR]

A thermophilic glycoside hydrolase family 16 (GH16) β-1,3-1,4-glucanase from Clostridium thermocellum (CtLic16A) holds great potentials in industrial applications due to its high specific activity and outstanding thermostability. In order to understand its molecular machinery, the crystal structure of CtLic16A was determined to 1.95 Å resolution. The enzyme folds into a classic GH16 β-jellyroll architecture which consists of two β-sheets atop each other, with the substrate-binding cleft lying on the concave side of the inner β-sheet. Two Bis–Tris propane molecules were found in the positive and negative substrate binding sites. Structural analysis suggests that the major differences between the CtLic16A and other GH16 β-1,3-1,4-glucanase structures occur at the protein exterior. Furthermore, the high catalytic efficacy and thermal profile of the CtLic16A are preserved in the enzyme produced in Pichia pastoris , encouraging its further commercial applications. [ABSTRACT FROM AUTHOR]

Abstract: Alkaline phytases from Bacillus species, which hydrolyze phytate to less phosphorylated myo-inositols and inorganic phosphate, have great potential as additives to animal feed. The thermostability and neutral optimum pH of Bacillus phytase are attributed largely to the presence of calcium ions. Nonetheless, no report has demonstrated directly how the metal ions coordinate phytase and its substrate to facilitate the catalytic reaction. In this study, the interactions between a phytate analog (myo-inositol hexasulfate) and divalent metal ions in Bacillus subtilis phytase were revealed by the crystal structure at 1.25 Å resolution. We found all, except the first, sulfates on the substrate analog have direct or indirect interactions with amino acid residues in the enzyme active site. The structures also unraveled two active site-associated metal ions that were not explored in earlier studies. Significantly, one metal ion could be crucial to substrate binding. In addition, binding of the fourth sulfate of the substrate analog to the active site appears to be stronger than that of the others. These results indicate that alkaline phytase starts by cleaving the fourth phosphate, instead of the third or the sixth that were proposed earlier. Our high-resolution, structural representation of Bacillus phytase in complex with a substrate analog and divalent metal ions provides new insight into the catalytic mechanism of alkaline phytases in general. [Copyright &y& Elsevier]