Addition of C-terminal histidyl tags to PsaL and PsaK1 proteins of cyanobacterial photosystem I.
Indian J Biochem Biophys
;
2000 Dec; 37(6): 433-40
Article
in English
| IMSEAR
| ID: sea-27015
ABSTRACT
In vitro mutagenesis was used to produce two photosystem I mutants of the cyanobacterium Synechocystis sp. PCC 6803. The mutant HK and HL contained hexahistidyl tags at the C-termini of the PsaK1 and PsaL subunits, respectively. The HK mutant contained wild-type amounts of trimeric PS I complexes, but the level of hexahistidine-tagged PsaK1 was found only ten per cent in the PS I complexes and membranes of the wild type level. Therefore, attachment of a tag at the C-terminus interferes with the expression or assembly of PsaK1. In contrast, the HL mutant contained a similar level of tagged PsaL as that in the wild type. However, trimeric PS I complexes could not be obtained from this strain, indicating that the C-terminus of PsaL is involved in the formation of PS I trimers. Hexahistidine-tagged complexes of the HL and HK strains could not be purified with Nickel-affinity chromatography, unless photosystem I was denatured with urea, demonstrating that tagged C-termini of PsaK1 and PsaL were embedded inside of the PS I complex. Protection of the C-terminus from trypsin cleavage further supported this conclusion. Thus, histidine tagging allowed us to demonstrate role of C-termini of two proteins of photosystem I.
Full text:
Available
Index:
IMSEAR (South-East Asia)
Main subject:
Base Sequence
/
Mutagenesis
/
Cyanobacteria
/
DNA Primers
/
Photosynthetic Reaction Center Complex Proteins
/
Photosystem I Protein Complex
/
Histidine
Language:
English
Journal:
Indian J Biochem Biophys
Year:
2000
Type:
Article
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