Biomaterial Database

Mutagenesis of the Lactobacillus casei folylpolyglutamate synthetase gene at essential residues resembling an ATP binding site. 1994

J Toy, and A L Bognar

Department of Microbiology, University of Toronto, Ontario, Canada.

Site-directed mutagenesis studies were performed on a region of the Lactobacillus casei folylpoly-gamma-glutamate synthetase (FPGS) protein (residues 49 to 52), which is highly conserved when compared to the Escherichia coli and human FPGS proteins. The amino acid sequence of this region, GKGS/T, is similar to the consensus sequence for the A region of a nucleotide binding site, a motif which encodes a phosphate-binding loop. Mutation G49A or K50R, with substitution to amino acids of similar size and charge, resulted in decreases in Vmax/Km of 40- to over 100-fold, depending on the variable substrate. Alteration of G51 to S or T resulted in a large increase in the Km for glutamate. The Km for ATP was not affected more than 4-fold by any of the mutations. Our studies indicate that the conserved region is essential for FPGS function, since many of the mutations resulting in functionally conservative substitutions produced inactive enzymes. However, the mutations affected binding of all three substrates, so there is no direct evidence for involvement of the region in ATP binding.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D007780	Lacticaseibacillus casei	A rod-shaped bacterium isolated from milk and cheese, dairy products and dairy environments, sour dough, cow dung, silage, and human mouth, human intestinal contents and stools, and the human vagina. L. casei is CATALASE positive.	Lactobacillus casei
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D009838	Oligodeoxyribonucleotides	A group of deoxyribonucleotides (up to 12) in which the phosphate residues of each deoxyribonucleotide act as bridges in forming diester linkages between the deoxyribose moieties.	Oligodeoxynucleotide,Oligodeoxyribonucleotide,Oligodeoxynucleotides
D010453	Peptide Synthases	Ligases that catalyze the joining of adjacent AMINO ACIDS by the formation of carbon-nitrogen bonds between their carboxylic acid groups and amine groups.	Peptide Synthetases,Acid-Amino-Acid Ligases,Acid Amino Acid Ligases,Ligases, Acid-Amino-Acid,Synthases, Peptide,Synthetases, Peptide
D011994	Recombinant Proteins	Proteins prepared by recombinant DNA technology.	Biosynthetic Protein,Biosynthetic Proteins,DNA Recombinant Proteins,Recombinant Protein,Proteins, Biosynthetic,Proteins, Recombinant DNA,DNA Proteins, Recombinant,Protein, Biosynthetic,Protein, Recombinant,Proteins, DNA Recombinant,Proteins, Recombinant,Recombinant DNA Proteins,Recombinant Proteins, DNA
D003001	Cloning, Molecular	The insertion of recombinant DNA molecules from prokaryotic and/or eukaryotic sources into a replicating vehicle, such as a plasmid or virus vector, and the introduction of the resultant hybrid molecules into recipient cells without altering the viability of those cells.	Molecular Cloning
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005798	Genes, Bacterial	The functional hereditary units of BACTERIA.	Bacterial Gene,Bacterial Genes,Gene, Bacterial
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man