title: Methods to Characterize Folding and Function of BamA Cross-Link Mutants.
abstract: The utility of protein engineering, both the mutation and deletion of specific amino acids, to investigate protein structure and function has been demonstrated time and time again, and intermolecular and intramolecular interactions within the BAM complex and its individual components are no exception. Extensive efforts have probed conserved and unique amino acid sequences of the Bam proteins to define their functional roles. This chapter summarizes efforts as applied to the disulfide cross-link mutants of BamA and describes experimental methods used in our studies to determine that lateral opening of the barrel domain is required for function.
authors.affiliation: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA.
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Kuszak
authors.affiliation: Department of Biological Sciences, Markey Center for Structural Biology, Purdue University, West Lafayette, IN, 47907, USA.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Noinaj
authors.affiliation: Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA. skbuchan@helix.nih.gov.
authors.forename: Susan K
authors.initials: SK
authors.lastname: Buchanan
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015153:Blotting, Western; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K / Q000378:metabolism; D006358:Hot Temperature; D016296:Mutagenesis; D009154:Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary
chemical_list: D001425:Bacterial Outer Membrane Proteins; D019286:Endopeptidase K
keywords: BAM complex; BamA; Disulfide cross-linking; Outer membrane protein; Proteinase K; β-barrel membrane protein