Key residues for membrane binding, oligomerization, and pore forming activity of staphylococcal alpha-hemolysin identified by cysteine scanning mutagenesis and targeted chemical modification.
AuthorsWalker B, Bayley H
JournalJ Biol Chem
PubMed ID7559447
'The alpha-hemolysin (alpha HL) polypeptide is secreted by Staphylococcus aureus as a water-soluble monomer that assembles into lipid bilayers to form cylindrical heptameric pores 1-2 nm in effective internal diameter. We have individually replaced each charged residue (79 of 293 amino acids) and four neutral residues in alpha HL with ... More
Subunit exchange, conformational stability, and chaperone-like function of the small heat shock protein 16.5 from Methanococcus jannaschii.
AuthorsBova MP, Huang Q, Ding L, Horwitz J
JournalJ Biol Chem
PubMed ID12176992
'Hsp16.5, isolated from the hyperthermophilic Archaea Methanococcus jannaschii, is a member of the small heat-shock protein family. Small Hsps have 12- to 42-kDa subunit sizes and have sequences that are conserved among all organisms. The recently determined crystal structure of Hsp16.5 indicates that it consists discretely of 24 identical subunits. ... More
Evidence for multiple mechanisms for membrane binding and integration via carboxyl-terminal insertion sequences.
'Subcellular localization of proteins with carboxyl-terminal insertion sequences requires the molecule be both targeted to and integrated into the correct membrane. The mechanism of membrane integration of cytochrome b5 has been shown to be promiscuous, spontaneous, nonsaturable, and independent of membrane proteins. Thus endoplasmic reticulum localization for cytochrome b5 depends ... More
Insight into the conformation of protein folding intermediate(s) trapped by GroEL.
AuthorsTorella C, Mattingly JR, Artigues A, Iriarte A, Martinez-Carrion M
JournalJ Biol Chem
PubMed ID9461576
'Many aspects of the mechanism by which the GroEL/ES chaperonins mediate protein folding are still unclear, including the amount of structure present in the substrate bound to GroEL. To address this issue we have analyzed the susceptibility to limited proteolysis and to alkylation of cysteine residues of mitochondrial aspartate aminotransferase ... More
Surface labeling of key residues during assembly of the transmembrane pore formed by staphylococcal alpha-hemolysin.
AuthorsKrishnasastry M, Walker B, Braha O, Bayley H
JournalFEBS Lett
PubMed ID7988723
'Structural changes in staphylococcal alpha-hemolysin (alpha HL) that occur during oligomerization and pore formation on membranes have been examined by using a simple gel-shift assay to determine the rate of modification of key single-cysteine mutants with the hydrophilic sulfhydryl reagent, 4-acetamido-4''-((iodoacetyl)amino)stilbene-2,2''-disulfonate (IASD). The central glycine-rich loop of alpha HL lines ... More
Oxidative stress and responses in Arabidopsis thaliana and Oryza sativa subjected to chilling and salinity stress.
Subunit stoichiometry of staphylococcal alpha-hemolysin in crystals and on membranes: a heptameric transmembrane pore.
AuthorsGouaux JE, Braha O, Hobaugh MR, Song L, Cheley S, Shustak C, Bayley H
JournalProc Natl Acad Sci U S A
PubMed ID7809129
Elucidation of the accurate subunit stoichiometry of oligomeric membrane proteins is fraught with complexities. The interpretations of chemical cross-linking, analytical ultracentrifugation, gel filtration, and low-resolution electron microscopy studies are often ambiguous. Staphylococcal alpha-hemolysin (alpha HL), a homooligomeric toxin that forms channels in cell membranes, was believed to possess six subunits ... More
Biochemical and genetic evidence for three transmembrane domains in the class I holin, lambda S.
AuthorsGründling A, Bläsi U, Young R
JournalJ Biol Chem
PubMed ID10625606
lambda S, the prototype class I holin gene, encodes three potential transmembrane domains in its 107 codons, whereas 21 S, the class II prototype spans only 71 codons and encodes two transmembrane domains. Many holin genes, including lambda S and 21 S, have the "dual-start" regulatory motif at the N ... More
Sulfhydryl chemistry detects three conformations of the lipid binding region of Escherichia coli pyruvate oxidase.
AuthorsChang YY, Cronan JE
JournalBiochemistry
PubMed ID9305946
Site-specific disulfide cross-linking experiments detected a conformational change within the C-terminal segment of Escherichia coli pyruvate oxidase (PoxB), a lipid-activated homotetrameric enzyme, upon substrate binding [Chang, Y.-Y., & Cronan, J. E., Jr. (1995) J. Biol. Chem. 270, 7896-7901]. The C-terminal lipid binding regions were cross-linked only in the presence of ... More
Exploring the membrane domain of the reduced nicotinamide adenine dinucleotide-quinone oxidoreductase of Paracoccus denitrificans: characterization of the NQO7 subunit.
AuthorsBernardo SD, Yano T, Yagi T
JournalBiochemistry
PubMed ID10924136
The proton-translocating reduced nicotinamide adenine dinucleotide- (NADH-) quinone oxidoreductase (NDH-1) of Paracoccus denitrificans is composed of at least 14 different subunits (NQO1-14). In addition, this enzyme complex houses one flavin mononucleotide (FMN) and 7-8 iron-sulfur clusters as cofactors. The expression and partial characterization of the NQO7 subunit, one of the ... More
Subunit exchange of alphaA-crystallin.
AuthorsBova MP, Ding LL, Horwitz J, Fung BK
JournalJ Biol Chem
PubMed ID9368012
alpha-Crystallin, the major protein in the mammalian lens, is a molecular chaperone that can bind denaturing proteins and prevent their aggregation. Like other structurally related small heat shock proteins, each alpha-crystallin molecule is composed of an average of 40 subunits that can undergo extensive reorganization. In this study we used ... More