Aspartate aminotransferase and tylosin biosynthesis in Streptomyces fradiae.
Aspartate aminotransferase as well as valine dehydrogenase and threonine dehydratase was required for the biosynthesis of tylosin in Streptomyces fradiae NRRL 2702. The biosynthesis of these enzymes and tylosin production were repressed by high concentrations of ammonium ions. The change in specific tylosin production rates in batch cultures with different initial concentrations of ammonium ions showed patterns similar to those of the specific production rates of aspartate aminotransferase, valine dehydrogenase, and threonine dehydratase. Aspartate aminotransferase has been purified by acetone precipitation, DEAE-cellulose, hydroxyapatite, and preparative electrophoresis chromatographies. The purified enzyme (120 kDa) consisted of two subunits identical in molecular mass (54 kDa) and showed homogeneity, giving one band with a pI of 4.2 upon preparative isoelectric focusing. The enzyme was specific for L-aspartate in the forward reaction; the Km values were determined to be 2.7 mM for L-aspartate, 0.7 mM for 2-oxyglutarate, 12.8 mM for L-glutamate, and 0.15 mM for oxaloacetate. The enzyme was somewhat thermostable, having a maximum activity at 55 degrees C, and had a broad pH optimum that ranged from 5.5 to 8.0. The mode of action was a ping-pong-bi-bi mechanism.
Lee
S H
SH
Department of Microbiology, College of Natural Sciences, Seoul, Korea.
Lee
K J
KJ
D000594:Amino Acid Oxidoreductases / Q000378:metabolism; D000641:Ammonia / Q000494:pharmacology; D001219:Aspartate Aminotransferases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005285:Fermentation; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007700:Kinetics; D008970:Molecular Weight; D013302:Streptomyces / Q000187:drug effects / Q000378:metabolism*; D013696:Temperature; D013913:Threonine Dehydratase / Q000378:metabolism; D015645:Tylosin / Q000096:biosynthesis*; D050802:Valine Dehydrogenase (NADP+)
D000641:Ammonia; D000594:Amino Acid Oxidoreductases; D050802:Valine Dehydrogenase (NADP+); D001219:Aspartate Aminotransferases; D013913:Threonine Dehydratase; D015645:Tylosin