Interaction between primary and secondary metabolism in Streptomyces coelicolor A3(2): Role of pyrroline-5-carboxylate dehydrogenase

Douglas Robert Drummond, S. Smith, N. J. Wood, D. A. Hodgson

Research output: Contribution to journalArticle

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Abstract

The activity of the proline catabolic enzyme pyrroline-5-carboxylate dehydrogenase (EC 1.5.1.12) was induced up to threehundredfold by the addition of three hundred proline to the growth medium of the Gram-positive bacterium Streptomyces coelicolor A3(2). Rifampicin, an inhibitor of RNA polymerase activity, abolished induction, implying that regulation was at the level of activation of gene transcription. The enzyme was purified and SDS-PAGE of the highly purified enzyme preparation revealed a single subunit with M(r) 68 000. A single band of protein, which also stained for enzyme activity, was observed after native gel electrophoresis. The M(r) of the enzyme was estimated to be approximately 265 000 by native gel electrophoresis and approximately 305 000 by gel filtration, which indicated that the enzyme had a tetrameric quaternary structure. The apparent K(m) for pyrroline-5-carboxylate was 109 ± 7.3 μM, whilst that for NAD+ was 43.3 ± 2.5 μM. Product inhibition by NADH (apparent K(i) 0.6 mM) was observed. The observed V(max) was 22.0 ± 1 mol min-1 (mg protein)-1. Neither 1 nor 5 mM proline had any effect on enzyme activity,, whilst glutamate was a very weak inhibitor.

Original languageEnglish
Pages (from-to)1739-1744
Number of pages6
JournalMicrobiology
Volume141
Issue number7
Publication statusPublished - Jan 1 1995

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Streptomyces coelicolor
Secondary Metabolism
Oxidoreductases
Enzymes
Proline
NAD
Transcriptional Activation
Electrophoresis
Gels
Gram-Positive Bacteria
DNA-Directed RNA Polymerases
Rifampin
delta-1-pyrroline-5-carboxylate
Gel Chromatography
Polyacrylamide Gel Electrophoresis
Glutamic Acid
Proteins
Growth

All Science Journal Classification (ASJC) codes

  • Microbiology

Cite this

Interaction between primary and secondary metabolism in Streptomyces coelicolor A3(2) : Role of pyrroline-5-carboxylate dehydrogenase. / Drummond, Douglas Robert; Smith, S.; Wood, N. J.; Hodgson, D. A.

In: Microbiology, Vol. 141, No. 7, 01.01.1995, p. 1739-1744.

Research output: Contribution to journalArticle

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