PubMed · 7686830
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M H Schreier. Immunophilins and immunosuppression.. https://pubmed.ncbi.nlm.nih.gov/7686830/
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The murI gene encoding D-glutamate racemase plays an important role in the biosynthesis of D-glutamic acid, an essential component of cell wall peptidoglycan of almost all eubacteria. A DNA fragment that could rescue the auxotrophy of D-glutamic acid in the Escherichia coli murI mutant strain WM335 was isolated from Brevibacterium lactofermentum ATCC 13869 belonging to the coryneform bacteria. DNA sequencing reveals that it encodes a protein of 284 amino acid residues, which shows a high level of homology with D-glutamate racemases from several other bacteria.
Extracellular levansucrase (LevU) and sucrase (InvB) are two of the three saccharolytic enzymes involved in the sucrose metabolism of Zymomonas mobilis. The levU and invB genes were clustered with a 155bp interval on the chromosome. Both genes were transcribed constitutively at the basal level and the transcription of both genes was induced significantly when sucrose was added to the medium. These genes were transcribed as a bicistronic mRNA and the expression was modulated by a single promoter, which is located upstream of the levU gene. The transcriptional initiation site was mapped to -64bp from the translation start site of levU gene. These results indicated that two genes are most likely to constitute an operon. The glk operon, which encodes four glycolytic enzymes, was located close to the levU operon on the chromosome. Two apparent ORFs (ORF3 and 4) were found at the intervening sequence located between the glk and levU operons. These ORFs were transcribed divergently and showed high homology at the amino acid level with the bacterial global regulatory protein (Lrp) and aspartate racemase.
Seven unique enzymatic steps lead to the biosynthesis of L-lysine from L-aspartate semialdehyde and pyruvate in bacteria. The immediate precursor to L-lysine is D,L-diaminopimelate, a diamino acid which is incorporated into the pentapeptide of the Gram-negative peptidoglycan moiety. D,L-Diaminopimelate is generated from the corresponding L,L-isomer by the dapF-encoded epimerase. Diaminopimelate epimerase is a representative of the pyridoxal phosphate-independent amino acid racemases, for which substantial evidence exists supporting the role of two cysteine residues as the catalytic acid and base. The pH dependencies of the maximum velocities in the L,L --> D,L and D,L --> L,L direction depend on a single catalytic group exhibiting pK values of 7.0 and 6.1, respectively, which must be unprotonated for activity. The pH dependencies of the V/K values in both directions depend on the ionization of two groups, one exhibiting a pK value of 6.7 which must be unprotonated and one exhibiting a pK value of 8.5 which must be protonated. Primary kinetic isotope effects on V and V/K are unequal, with D(V/K) being larger than DV in both the forward and reverse directions. Solvent kinetic isotope effects in both directions are inverse on V/K, but normal on V. Both of these isotopic observations support a model in which proton isomerization after catalysis and substrate dissociation is kinetically significant. A single solvent "overshoot" is observed when L, L-diaminopimelate is incubated with enzyme in D2O; however, an unprecedented double overshoot is observed when D,L-diaminopimelate is incubated with enzyme in D2O. A model has been developed which allows these two overshoots to be simulated. A chemical mechanism is proposed invoking the function of two cysteine residues, Cys73 and Cys217, observed in the recently determined three-dimensional structure of this enzyme [Cirilli, M., et al. (1998) Biochemistry 37, 16452-16458], as the acid and base in the mechanism.