Elucidation of evolutionary factors that enhance protein thermostability is a critical problem and was the focus of this work on Thermus species. Pairs of orthologous sequences of T. scotoductus SA-01 and T. thermophilus HB27, with the largest negative minimum folding energy (MFE) as predicted by the UNAFold algorithm, were statistically analyzed. Favored substitutions of amino acids residues and their properties were determined. Substitutions were analyzed in modeled protein structures to determine their locations and contribution to energy differences using PyMOL and FoldX programs respectively. Dominant trends in amino acid substitutions consistent with differences in thermostability between orthologous sequences were observed. T. thermophilus thermophilic proteins showed an increase in non-polar, tiny, and charged amino acids. An abundance of alanine substituted by serine and threonine, as well as arginine substituted by glutamine and lysine was observed in T. thermophilus HB27. Structural comparison showed that stabilizing mutations occurred on surfaces and loops in protein structures.
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http://dx.doi.org/10.4137/EBO.S12539 | DOI Listing |
Sci Rep
January 2025
Laboratory of Extremophiles Biology, Department of Microbiology, Faculty of Biology, University of Gdansk, Wita Stwosza 59, Gdansk, 80-308, Poland.
In this study, we evaluated the combined effect between MLE-15, a modular lytic enzyme composed of four building blocks, and reline, a natural deep eutectic solvent. The bioinformatic analysis allowed us to determine the spatial architecture of MLE-15, whose components were bactericidal peptide cecropin A connected via a flexible linker to the cell wall binding domain (CBD) of mesophilic 201ϕ2 - 1 endolysin and catalytic domain (EAD) of highly thermostable Ph2119 endolysin. The modular enzyme showed high thermostability with the melting temperature of 93.
View Article and Find Full Text PDFVet Res
January 2025
Key Laboratory of Animal Genetics, Breeding and Reproduction in The Plateau Mountainous Region, Ministry of Education, Guizhou University, Guiyang, 550025, China.
Newcastle disease virus (NDV) is a significant member of the Paramyxoviridae family, known for causing epidemics and substantial economic losses in the poultry industry worldwide. The NDV RNA genome primarily encodes six structural proteins (N, P, M, F, HN, and L) and two non-structural proteins (V and W). Among these, the polymerase-associated proteins (N, P, and L) and the viral RNA (vRNA) genome form the ribonucleoprotein complex, which plays a crucial role in the synthesis and transcription of NDV vRNA.
View Article and Find Full Text PDFBioorg Chem
January 2025
Department of Bioengineering, Gebze Technical University, 41400, Gebze, Kocaeli, Türkiye; BAUZYME Biotechnology Co., Gebze Technical University Technopark, 41400, Gebze, Kocaeli, Türkiye. Electronic address:
α-Amylases, constituting a significant share of the enzyme market, are mainly synthesized by the genus Bacillus. Enzymes tailored for specific industrial applications are needed to meet the growing demand across a range of industries, and thus finding new amylases and optimizing the ones that already exist are extremely important. This study reports the successful expression, characterization and immobilization of P.
View Article and Find Full Text PDFSTAR Protoc
January 2025
Department of Molecular Medicine, University of Pavia, Pavia, Italy. Electronic address:
Voltage-dependent anion channel 1 (VDAC1) is a key protein in cellular metabolism and apoptosis. Here, we present a protocol to express and purify milligram amounts of recombinant VDAC1 in Escherichia coli. We detail steps for a fluorescence polarization-based high-throughput screening assay using NADH displacement, along with procedures for thermostability, fluorescence polarization, and X-ray crystallography.
View Article and Find Full Text PDFNat Commun
January 2025
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, PR China.
The pursuit of obtaining enzymes with high activity and stability remains a grail in enzyme evolution due to the stability-activity trade-off. Here, we develop an isothermal compressibility-assisted dynamic squeezing index perturbation engineering (iCASE) strategy to construct hierarchical modular networks for enzymes of varying complexity. Molecular mechanism analysis elucidates that the peak of adaptive evolution is reached through a structural response mechanism among variants.
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