Structure and expression of the Chlorobium vibrioforme hemA gene

Abstract

The green sulfur bacterium, Chlorobium vibrioforme, synthesizes the tetrapyrrole precursor, delta-aminolevulinic acid (ALA), from glutamate via the RNA-dependent five-carbon pathway. A 1.9-kb clone of genomic DNA from C. vibrioforme that is capable of transforming a glutamyl-tRNA reductase-deficient, ALA-dependent, hemA mutant of Escherichia coli to prototrophy was sequenced. The transforming C. vibrioforme DNA has significant sequence similarity to the E. coli, Salmonella typhimurium, and Bacillus subtilis hemA genes and contains a 1245 base open reading frame that encodes a 415 amino acid polypeptide with a calculated molecular weight of 46174. This polypeptide has over 28% amino acid identity with the polypeptides deduced from the nucleic acid sequences of the E. coli, S. typhimurium, and B. subtilis hemA genes. No sequence similarity was detected, at either the nucleic acid or the peptide level, with the Rhodobacter capsulatus or Bradyrhizobium japonicum hemA genes, which encode ALA synthase, or with the S. typhimurium hemL gene, which encodes glutamate-1-semialdehyde aminotransferase. These results establish that hemA encodes glutamyl-tRNA reductase in species that use the five-carbon ALA biosynthetic pathway. A second region of the cloned DNA, located downstream from the hemA gene, has significant sequence similarity to the E. coli and B. subtilis hemC genes. This region contains a potential open reading frame that encodes a polypeptide that has high sequence identity to the deduced E. coli and B. subtilis HemC peptides. hemC encodes the tetrapyrrole biosynthetic enzyme, porphobilinogen deaminase, in these species.(ABSTRACT TRUNCATED AT 250 WORDS)

Purification and partial characterisation of barley glutamyl‐tRNAGlu reductase, the enzyme that directs glutamate to chlorophyll biosynthesis

“…This suggestion was supported by the finding that complementation of the h e 4 mutation in E. coli by a cloned DNA fragment from Chlorobium vibrioforme changed the tRNA specificity of the restored 5-aminolevulinic-acid-synthesizing activity to resemble that of Chlorobium. The cloned DNA fragment was later sequenced, and shown to have an open reading frame which at the amino acid level showed 28.4% identity with the E. coli hemA gene product (Majumdar et al, 1991). Complementation cloning has been used to isolate the hemA gene from Salmonella typhimurium (Elliott, 1989), Bacillus subtilis (Petricek et al, 1990), Synechocystis (Grimm, 1992;Verkamp et al, 1992), Arabidopsis thaliana (Ilag et al, 1994), Coxiella burnetii (EMBL X78969), Clostridium josui (EMBL D28503) and barley (Grimm, B. and Bougri, O., unpublished results).…”

“…(Ilaget al, 1994); synts, Synechocystis (Grimm, 1992); ecoli, E. coli (Verkamp and Chelm, 1989); salty, S. Qphimuriurn (Elliott, 1989); coxbr, Coxiellu burnetii (Willems et al, 1994, EMBL database); chvib, C. vibriofonne (Majumdar et al, 1991); cljoj, C. josui (Fujino et al, 1994, EMBL database); and conse, consensus sequence. Amino acids are included in the consensus sequence, if they occur in six or more of the aligned sequences, or if the changes are considered conservative.…”

Functional analysis of the promoter region of the human phosphotyrosine phosphatase activator gene: Yin Yang 1 is essential for core promoter activity

“…High-quality genomic DNA was isolated from liver HepG2 cells as described by Majumdar et al [17]. A 30 µg sample of DNA was double-digested with SacI\NaeI, SacI\NarI or SacI\EagI, or with SacI alone as a control.…”

Cloning and characterization of the Bacillus subtilis hemEHY gene cluster, which encodes protoheme IX biosynthetic enzymes

“…Genetic loci required for protoheme IX synthesis are tightly linked on the S. aureus chromosome (49). The hem genes of gram-negative bacteria in contrast are scattered on the chromosome, with the exception of the hemCD operon of E. coli (19,44) and Salmonella typhimuium (50) and a possible hemACD operon in Chlorobium vibrioforme (24).…”