Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.

A reaction center H- strain (RCH-) of Rhodobacter sphaeroides, PUHA1, was made by in vitro deletion of an XhoI restriction endonuclease fragment from the puhA gene coupled with insertion of a kanamycin resistance gene cartridge. The resulting construct was delivered to R. sphaeroides wild-type 2.4.1...

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Main Authors: Sockett, R E, Donohue, T J, Varga, A R, Kaplan, S
Formato: Artigo
Idioma:en
Publicado em: 1989
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Research Article
Acesso em linha:https://ncbi.nlm.nih.gov/pmc/articles/PMC209607/
https://ncbi.nlm.nih.gov/pubmed/2644200
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spelling pubmed-2096072003-12-01 Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences. Sockett, R E Donohue, T J Varga, A R Kaplan, S J Bacteriol Research Article A reaction center H- strain (RCH-) of Rhodobacter sphaeroides, PUHA1, was made by in vitro deletion of an XhoI restriction endonuclease fragment from the puhA gene coupled with insertion of a kanamycin resistance gene cartridge. The resulting construct was delivered to R. sphaeroides wild-type 2.4.1, with the defective puhA gene replacing the wild-type copy by recombination, followed by selection for kanamycin resistance. When grown under conditions known to induce intracytoplasmic membrane development, PUHA1 synthesized a pigmented intracytoplasmic membrane. Spectral analysis of this membrane showed that it was deficient in B875 spectral complexes as well as functional reaction centers and that the level of B800-850 spectral complexes was greater than in the wild type. The RCH- strain was photosythetically incompetent, but photosynthetic growth was restored by complementation with a 1.45-kilobase (kb) BamHI restriction endonuclease fragment containing the puhA gene carried in trans on plasmid pRK404. B875 spectral complexes were not restored by complementation with the 1.45-kb BamHI restriction endonuclease fragment containing the puhA gene but were restored along with photosynthetic competence by complementation with DNA from a cosmid carrying the puhA gene, as well as a flanking DNA sequence. Interestingly, B875 spectral complexes, but not photosynthetic competence, were restored to PUHA1 by introduction in trans of a 13-kb BamHI restriction endonuclease fragment carrying genes encoding the puf operon region of the DNA. The effect of the puhA deletion was further investigated by an examination of the levels of specific mRNA species derived from the puf and puc operons, as well as by determinations of the relative abundances of polypeptides associated with various spectral complexes by immunological methods. The roles of puhA and other genetic components in photosynthetic gene expression and membrane assembly are discussed. 1989-01 /pmc/articles/PMC209607/ /pubmed/2644200 Text en
institution US National Library of Medicine
collection PubMed Central
language en
format Article
topic Research Article
spellingShingle Research Article
Sockett, R E
Donohue, T J
Varga, A R
Kaplan, S
Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
description A reaction center H- strain (RCH-) of Rhodobacter sphaeroides, PUHA1, was made by in vitro deletion of an XhoI restriction endonuclease fragment from the puhA gene coupled with insertion of a kanamycin resistance gene cartridge. The resulting construct was delivered to R. sphaeroides wild-type 2.4.1, with the defective puhA gene replacing the wild-type copy by recombination, followed by selection for kanamycin resistance. When grown under conditions known to induce intracytoplasmic membrane development, PUHA1 synthesized a pigmented intracytoplasmic membrane. Spectral analysis of this membrane showed that it was deficient in B875 spectral complexes as well as functional reaction centers and that the level of B800-850 spectral complexes was greater than in the wild type. The RCH- strain was photosythetically incompetent, but photosynthetic growth was restored by complementation with a 1.45-kilobase (kb) BamHI restriction endonuclease fragment containing the puhA gene carried in trans on plasmid pRK404. B875 spectral complexes were not restored by complementation with the 1.45-kb BamHI restriction endonuclease fragment containing the puhA gene but were restored along with photosynthetic competence by complementation with DNA from a cosmid carrying the puhA gene, as well as a flanking DNA sequence. Interestingly, B875 spectral complexes, but not photosynthetic competence, were restored to PUHA1 by introduction in trans of a 13-kb BamHI restriction endonuclease fragment carrying genes encoding the puf operon region of the DNA. The effect of the puhA deletion was further investigated by an examination of the levels of specific mRNA species derived from the puf and puc operons, as well as by determinations of the relative abundances of polypeptides associated with various spectral complexes by immunological methods. The roles of puhA and other genetic components in photosynthetic gene expression and membrane assembly are discussed.
author Sockett, R E
Donohue, T J
Varga, A R
Kaplan, S
author_facet Sockett, R E
Donohue, T J
Varga, A R
Kaplan, S
author_sort Sockett, R E
title Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
title_short Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
title_full Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
title_fullStr Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
title_full_unstemmed Control of photosynthetic membrane assembly in Rhodobacter sphaeroides mediated by puhA and flanking sequences.
title_sort control of photosynthetic membrane assembly in rhodobacter sphaeroides mediated by puha and flanking sequences.
publishDate 1989
url https://ncbi.nlm.nih.gov/pmc/articles/PMC209607/
https://ncbi.nlm.nih.gov/pubmed/2644200
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