Substrate Binding Properties of Mutant and Wild-Type A Proteins of Escherichia coli Tryptophan Synthetase

Science ◽  
1967 ◽  
Vol 156 (3780) ◽  
pp. 1369-1371 ◽  
Author(s):  
J. K. Hardman ◽  
C. Yanofsky
Keyword(s):  
Escherichia Coli ◽  
Substrate Binding ◽  
Type A ◽  
Wild Type ◽  
Binding Properties ◽  
Tryptophan Synthetase

scholarly journals Dependence on pH of substrate binding to a mutant lactose carrier, lacYun, in Escherichia coli. A model for H+/lactose symport

1989 ◽  
Vol 258 (2) ◽  
pp. 389-396 ◽  
Author(s):  
I Yamato ◽  
Y Anraku
Keyword(s):  
Escherichia Coli ◽  
Substrate Binding ◽  
Ph Dependence ◽  
Plasmid Vector ◽  
Binary Complex ◽  
Wild Type ◽  
Transport Reaction ◽  
Substrate Transport ◽  
Vector Pbr322 ◽  
Dissociation Step

The lacYun gene, which encodes a lactose carrier showing the uncoupled phenotype of substrate transport in Escherichia coli [Wilson, Kusch & Kashket (1970) Biochem. Biophys. Res. Commun. 40, 1409-1414], was cloned on a plasmid vector, pBR322. The binding of a substrate, p-nitrophenyl alpha-galactoside, to the lacYun carrier in membranes from the strain harbouring the lacYun clone showed a pH-dependence different from its binding to the wild-type lactose carrier. This finding indicated that the lacYun mutation confers higher affinity for H+ on the carrier, exerting its effect on the less efficient dissociation of substrate inside cells. The result coincides with the proposal [Yamato & Rosenbusch (1983) FEBS Lett. 151, 102-104] that the proton affecting the substrate binding is the coupling proton of the proton/lactose symport reaction, which allows only the ordered mechanism of binding of substrate to an H+-carrier binary complex. From the simplest model of the symport reaction, constructed on the basis of these results, the coupling site of energy in the carrier cycle of the transport reaction can be identified at the substrate-dissociation step inside cells.

scholarly journals Protein Synthesis in Escherichia coli with Mischarged tRNA

2003 ◽  
Vol 185 (12) ◽  
pp. 3524-3526 ◽  
Author(s):  
Bokkee Min ◽  
Makoto Kitabatake ◽  
Carla Polycarpo ◽  
Joanne Pelaschier ◽  
Gregory Raczniak ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Elongation Factor ◽  
Trna Synthetase ◽  
Wild Type ◽  
Wild Type Level ◽  
E Coli ◽  
Missense Mutant ◽  
Tryptophan Synthetase

ABSTRACT Two types of aspartyl-tRNA synthetase exist: the discriminating enzyme (D-AspRS) forms only Asp-tRNAAsp, while the nondiscriminating one (ND-AspRS) also synthesizes Asp-tRNAAsn, a required intermediate in protein synthesis in many organisms (but not in Escherichia coli). On the basis of the E. coli trpA34 missense mutant transformed with heterologous ND-aspS genes, we developed a system with which to measure the in vivo formation of Asp-tRNAAsn and its acceptance by elongation factor EF-Tu. While large amounts of Asp-tRNAAsn are detrimental to E. coli, smaller amounts support protein synthesis and allow the formation of up to 38% of the wild-type level of missense-suppressed tryptophan synthetase.

scholarly journals Lysine Residue 117 of the FasG Adhesin of Enterotoxigenic Escherichia coli Is Essential for Binding of 987P Fimbriae to Sulfatide

1999 ◽  
Vol 67 (11) ◽  
pp. 5755-5761 ◽  
Author(s):  
Byung-Kwon Choi ◽  
Dieter M. Schifferli
Keyword(s):  
Escherichia Coli ◽  
Essential Role ◽  
Surface Display ◽  
Salt Bridge ◽  
Site Directed Mutagenesis ◽  
Enterotoxigenic Escherichia Coli ◽  
Wild Type ◽  
Binding Properties ◽  
Brush Borders ◽  
Positively Charged

ABSTRACT The FasG subunit of the 987P fimbriae of enterotoxigenic strains ofEscherichia coli was previously shown to mediate fimbrial binding to a glycoprotein and a sulfatide receptor on intestinal brush borders of piglets. Moreover, the 987P adhesin FasG is required for fimbrial expression, since fasG null mutants are nonfimbriated. In this study, fasG was modified by site-directed mutagenesis to study its sulfatide binding properties. Twenty single mutants were generated by replacing positively charged lysine (K) or arginine (R) residues with small, nonpolar alanine (A) residues. Reduced levels of binding to sulfatide-containing liposomes correlated with reduced fimbriation and FasG surface display in fourfasG mutants (R27A, R286A, R226A, and R368). Among the 16 remaining normally fimbriated mutants with wild-type levels of surface-exposed FasG, only one mutant (K117A) did not interact at all with sulfatide-containing liposomes. Four mutants (K117A, R116A, K118A, and R200A) demonstrated reduced binding to such liposomes. Since complete phenotypic dissociation between the structure and specific function of 987P was observed only with mutant K117A, this residue is proposed to play an essential role in the FasG-sulfatide interaction, possibly communicating with the sulfate group of sulfatide by hydrogen bonding and/or salt bridge formation. Residues K17, R116, K118, and R200 may stabilize this interaction.

scholarly journals YfiK from Escherichia coli Promotes Export of O-Acetylserine and Cysteine

2003 ◽  
Vol 185 (4) ◽  
pp. 1161-1166 ◽  
Author(s):  
Isabel Franke ◽  
Armin Resch ◽  
Tobias Daßler ◽  
Thomas Maier ◽  
August Böck
Keyword(s):  
Escherichia Coli ◽  
Membrane Protein ◽  
Gene Product ◽  
Double Mutant ◽  
Type A ◽  
Integral Membrane Protein ◽  
Transmembrane Helices ◽  
Wild Type ◽  
In The Wild ◽  
Production Strain

ABSTRACT yfiK was discovered as a gene augmenting cysteine production when it was overexpressed in an industrial Escherichia coli production strain. The gene product is an integral membrane protein with about six predicted transmembrane helices; it belongs to the RhtB family of export proteins. YfiK overproduction from a plasmid leads to drastic and parallel secretion of O-acetylserine and cysteine into the medium but only when the organism possesses a serine transacetylase that is feedback insensitive to cysteine. Externally provided O-acetylserine obviated this requirement for cysteine secretion both in the yfiK-carrying transformant and in the wild type. A ΔyfiK mutant did not show any phenotype, and it exported O-acetylserine and cysteine when transformed with a plasmid carrying ydeD, a previously characterized, alternate O-acetylserine/cysteine exporter. Since a ydeD-yfiK double mutant showed the same pattern, it appears that YfiK and YdeD act independently. The necessity for the cell to regulate the size of the internal pool of O-acetylserine via synthesis of exporter proteins could be connected to the fact that this compound (when supplied externally) inhibits growth. Overexpression of either ydeD or yfiK leads to alleviation of this inhibition paralled by increased resistance to azaserine, which is an analog of O-acetylserine.

scholarly journals Tir Tyrosine Phosphorylation and Pedestal Formation Are Delayed in Enteropathogenic Escherichia coli sepZ::TnphoA Mutant 30-5-1(3)

2001 ◽  
Vol 69 (1) ◽  
pp. 559-563 ◽  
Author(s):  
Rebekah Devinney ◽  
Israel Nisan ◽  
Sharon Ruschkowski ◽  
Ilan Rosenshine ◽  
B. Brett Finlay
Keyword(s):  
Escherichia Coli ◽  
Tyrosine Phosphorylation ◽  
Type A ◽  
Secreted Proteins ◽  
Enteropathogenic Escherichia Coli ◽  
Wild Type ◽  
Epec Strain ◽  
Lesion Formation ◽  
Transposon Insertion ◽  
Pedestal Formation

ABSTRACT Enteropathogenic Escherichia coli (EPEC) strain 30-5-1(3) has been reported to form attaching and effacing (A/E) lesions without Tir tyrosine phosphorylation. In this study, we show that 30-5-1(3), which has a transposon insertion within thesepZ gene, forms wild-type A/E lesions including Tir tyrosine phosphorylation, but at a slower rate. A/E lesion formation by 30-5-1(3) occurs without detectable secretion of Tir or other EPEC Esp secreted proteins.

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