Expression of the Rhodobacter sphaeroides cytochrome c2 structural gene

J Bacteriol. 1989 Jan;171(1):360-8. doi: 10.1128/jb.171.1.360-368.1989.

Abstract

A Rhodobacter sphaeroides mutant (CYCA1) lacking cytochrome c2 (cyt c2) was previously constructed (T. J. Donohue, A. G. McEwan, S. Van Doren, A. R. Crofts, and S. Kaplan, Biochemistry, 27: 1918-1924, 1988) by a combination of in vivo and in vitro molecular genetic techniques. CYCA1 was incapable of photosynthetic growth (PS-); in this presentation, we show that chemoheterotrophically grown CYCA1 contained significant quantities of a high potential soluble c-type cytochrome(s) with an alpha band of approximately 554 nm which had previously gone undetected under these physiological conditions in wild-type cells. In addition, the PS- phenotype of CYCA1 can be complemented in trans with stable low-copy-number (approximately 5 to 9 per R. sphaeroides genome) broad-host-range plasmids containing the wild-type cyt c2 structural gene (cycA) and upstream regulatory sequences. cyt c2 and cycA-specific mRNA levels were elevated in both the wild type and CYCA1 derivatives harboring intact cycA genes in trans, presumably as a result of increased gene dosage. Although photosynthetically grown wild-type cells contained approximately twofold more cycA-specific transcripts than chemoheterotrophically grown cells, there was an approximately four- to sevenfold increase in cyt c2 levels under photosynthetic conditions. Similarly, complemented CYCA1 strains contained between 1.3- and 2.3-fold more cycA mRNA under photosynthetic conditions than under chemoheterotrophic conditions and had 6- to 12-fold higher steady-state levels of cyt c2 under the same physiological conditions. These data are discussed in terms of possible posttranscriptional control over cyt c2.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Cloning, Molecular
  • Cytochrome c Group / biosynthesis
  • Cytochrome c Group / genetics*
  • Cytochromes c2
  • DNA, Recombinant / metabolism
  • Escherichia coli / genetics
  • Genes*
  • Genes, Bacterial*
  • Genetic Complementation Test
  • RNA, Messenger / genetics
  • Restriction Mapping
  • Rhodobacter sphaeroides / genetics*
  • Rhodobacter sphaeroides / metabolism
  • Transcription, Genetic

Substances

  • Cytochrome c Group
  • DNA, Recombinant
  • RNA, Messenger
  • Cytochromes c2