title: Rapid Bioinformatic Identification of Thermostabilizing Mutations.
abstract: Ex vivo stability is a valuable protein characteristic but is laborious to improve experimentally. In addition to biopharmaceutical and industrial applications, stable protein is important for biochemical and structural studies. Taking advantage of the large number of available genomic sequences and growth temperature data, we present two bioinformatic methods to identify a limited set of amino acids or positions that likely underlie thermostability. Because these methods allow thousands of homologs to be examined in silico, they have the advantage of providing both speed and statistical power. Using these methods, we introduced, via mutation, amino acids from thermoadapted homologs into an exemplar mesophilic membrane protein, and demonstrated significantly increased thermostability while preserving protein activity.
authors.affiliation: Department of Cell Biology, The Helen L. and Martin S. Kimmel Center for Biology and Medicine, Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York. Electronic address: david.sauer@med.nyu.edu.
authors.forename: David B
authors.initials: DB
authors.lastname: Sauer
authors.affiliation: Department of Cell Biology, The Helen L. and Martin S. Kimmel Center for Biology and Medicine, Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York.
authors.forename: Nathan K
authors.initials: NK
authors.lastname: Karpowich
authors.affiliation: Department of Cell Biology, The Helen L. and Martin S. Kimmel Center for Biology and Medicine, Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York.
authors.forename: Jin Mei
authors.initials: JM
authors.lastname: Song
authors.affiliation: Department of Cell Biology, The Helen L. and Martin S. Kimmel Center for Biology and Medicine, Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, New York. Electronic address: da-neng.wang@med.nyu.edu.
authors.forename: Da-Neng
authors.initials: DN
authors.lastname: Wang
mesh_terms: D000596:Amino Acids / Q000737:chemistry / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D017920:Antiporters / Q000737:chemistry / Q000235:genetics; D001412:Bacillus subtilis; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D019295:Computational Biology / Q000379:methods*; D004926:Escherichia coli; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D055550:Protein Stability; D013696:Temperature; D013752:Tetracycline / Q000494:pharmacology; D014162:Transfection
chemical_list: D000596:Amino Acids; D000900:Anti-Bacterial Agents; D017920:Antiporters; D001426:Bacterial Proteins; C482657:TetL protein, Bacillus subtilis; D049452:Green Fluorescent Proteins; D013752:Tetracycline