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Selecting thioredoxins for disulphide proteomics: Target proteomes of three thioredoxins from the cyanobacterium Synechocystis sp. PCC 6803
Author(s) -
PérezPérez María Esther,
Florencio Francisco J.,
Lindahl Marika
Publication year - 2006
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200500215
Subject(s) - proteome , thioredoxin , biochemistry , proteomics , cysteine , enzyme , biology , serine , glycogen phosphorylase , synechocystis , active site , chemistry , mutant , gene
Abstract Searching for enzymes and other proteins which can be redox‐regulated by dithiol/disulphide exchange is a rapidly expanding area of functional proteomics. Recently, several experimental approaches using thioredoxins have been developed for this purpose. Thioredoxins comprise a large family of redox‐active enzymes capable of reducing protein disulphides to cysteines and of participating in a variety of processes, such as enzyme modulation, donation of reducing equivalents and signal transduction. In this study we screened the target proteomes of three different thioredoxins from the unicellular cyanobacterium Synechocystis sp. PCC 6803, using site‐directed active‐site cysteine‐to‐serine mutants of its m ‐, x ‐ and y ‐type thioredoxins. The properties of a thioredoxin that determine the outcome of such analyses were found to be target‐binding capacity, solubility and the presence of non‐active‐site cysteines. Thus, we explored how the choice of thioredoxin affects the target proteomes and we conclude that the m ‐type thioredoxin, TrxA, is by far the most useful for screening of disulphide proteomes. Furthermore, we improved the resolution of target proteins on non‐reducing/reducing 2‐DE, leading to the identification of 14 new potentially redox‐regulated proteins in this organism. The presence of glycogen phosphorylase among the newly identified targets suggests that glycogen breakdown is redox‐regulated in addition to glycogen synthesis.