1 Enzyme and Protein Chemistry, Department of Systems Biology, Technical University of Denmark2 Department of Systems Biology, Technical University of Denmark3 Enzyme and Protein Chemistry, Department of Systems Biology, Technical University of Denmark4 Center for Biological Sequence Analysis, Department of Systems Biology, Technical University of Denmark5 Department of Mechanical Engineering, Technical University of Denmark
Three protein disulfide reductases of the thioredoxin superfamily from the industrially important Gram-positive Lactococcus lactis (LlTrxA, LlTrxD and LlNrdH) are compared to the "classical" thioredoxin from Escherichia coil (EcTrx1). LlTrxA resembles EcTrx1 with a WCGPC active site motif and other key residues conserved. By contrast, LlTrxD is more distantly related and contains a WCGDC motif. Bioinformatics analysis suggests that LlTrxD represents a subgroup of thioredoxins from Gram-positive bacteria. LlNrdH is a glutaredoxin-like electron donor for ribonucleotide reductase class Ib. Based on protein-protein equilibria LlTrxA (E01 = -259 mV) and LlNrdH (E01 = -238 my) show approximately 10 mV higher standard state redox potentials than the corresponding E. coil homologues, while E 01 of LlTrxD is -243 mV, more similar to glutaredoxin than "classical" thioredoxin. EcTrx1 and LlTrxA have high capacity to reduce insulin disulfides and their exposed active site thiol is alkylated at a similar rate at pH 7.0. LlTrxD on the other hand, is alkylated by iodoacetamide at almost 100 fold higher rate and shows no activity towards insulin disulfides. LlTrxA, LlTrxD and L1NrdH are all efficiently reduced by NADPH dependent thioredoxin reductase (TrxR) from L lactis and good cross-reactivity towards E. coil TrxR was observed with LITrxD as the notable exception.
Archives of Biochemistry and Biophysics, 2014, Vol 564, p. 164-172