Pentalenolactone-insensitive glyceraldehyde-3-phosphate dehydrogenase from Streptomyces arenae is closely related to GAPDH from thermostable eubacteria and plant chloroplasts.
Streptomyces arenae produces the antibiotic pentalenolactone, a highly specific inhibitor of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). During the phase of pentalenolactone production, S. arenae expresses a pentalenolactone-insensitive GAPDH isoform; otherwise, a pentalenolactone-sensitive form is expressed. The gene of the pentalenolactone-insensitive GAPDH was cloned and sequenced. Regulatory elements typical for genes encoding antibiotic resistance and production are localized upstream and downstream of the open reading frame. No expression of pentalenolactone-insensitive GAPDH was detected in Streptomyces lividans transformed with the gene. In Escherichia coli, the gene was expressed from an induced lac promoter. Amino-terminal sequencing of the heterologously expressed GAPDH proved its identity with pentalenolactone-insensitive GAPDH from S. arenae. Sequence comparisons with GAPDH from other organisms showed a close relationship to GAPDH of plant chloroplasts, of other gram-positive bacteria, and of thermophilic gram-negative bacteria. Pentalenolactone-insensitive GAPDH differs from all closely related GAPDHs only in a few residues, none of which are directly involved in catalysis or substrate binding. The total amino acid composition is more similar to GAPDH of thermophilic species than to that of mesophilic species. The purified enzyme was moderately thermotolerant, which could be a side effect of the structural changes causing pentalenolactone-resistance.
Fröhlich
K U
KU
Physiologisch-chemisches Institut, Universität Tübingen, Hoppe-Seyler-Strasse 4, D-72076 Tübingen, Germany.
Kannwischer
R
R
Rüdiger
M
M
Mecke
D
D
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001419:Bacteria / Q000201:enzymology; D001482:Base Composition; D001483:Base Sequence; D002736:Chloroplasts / Q000201:enzymology; D003001:Cloning, Molecular; D004352:Drug Resistance, Microbial; D004795:Enzyme Stability; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D010944:Plants / Q000201:enzymology; D017422:Sequence Analysis, DNA; D012717:Sesquiterpenes / Q000494:pharmacology; D013302:Streptomyces / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D013696:Temperature
D000900:Anti-Bacterial Agents; D012717:Sesquiterpenes; C017996:arenaemycin E; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases