Affiliation:
1. Lehrstuhl für Allgemeine Mikrobiologie und Mikrobengenetik, FSU Jena, Neugasse 24, 07743 Jena, Germany
2. Leibniz-Institut für Naturstoff-Forschung und Infektionsbiologie—Hans-Knöll-Institut, Beutenbergstraße 11a, 07745 Jena, Germany
Abstract
ABSTRACT
The NADP-dependent 4-dihydrotrisporin-dehydrogenase is a (−) mating-type-specific enzyme in the pathway from β-carotene to trisporic acid. This substance and its isomers and derivatives represent the general system of sexual communication in zygomycetes. The (−) mating type of
Mucor mucedo
was stimulated by trisporic acid and the enzyme was purified by ion exchange and affinity chromatography. Several peptides of the 26-kDa protein, digested with trypsin, were sequenced by mass spectrometry. Oligonucleotides based on protein sequence data were used for PCR amplification of genomic DNA. The primary PCR fragment was sequenced and the complete gene,
TSP2
, was isolated. A labeled
TSP2
hybridization probe detects a single-copy gene in the genome of
M. mucedo
. Northern blot analysis with RNAs from different growth stages reveals that the expression of the gene depends on the developmental stage of the mycelium in both mating types of
M. mucedo
. At the enzyme level, activity is found exclusively in the (−) mating type. However, renaturation of proteins in sodium dodecyl sulfate-containing gels revealed the
TSP2
gene product in both mating types. Analyzing the protein sequence places the enzyme in the short chain dehydrogenase superfamily. Thus, it has an evolutionary origin distinct from that of the previously isolated 4-dihydromethyltrisporate dehydrogenase, which belongs to the aldo/keto reductase superfamily. Apart from the
TSP2
genes in the three sequenced zygomycetous genomes (
Phycomyces blakesleeanus
,
Rhizopus oryzae
, and
Mucor circinelloides
), the closest relative is the
Myxococcus xanthus CsgA
gene product, which is also a short chain dehydrogenase, involved in C signaling and fruiting body formation.
Publisher
American Society for Microbiology
Subject
Molecular Biology,General Medicine,Microbiology
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