REPLACEMENT OF PRO(109) BY HIS IN TLPA, A THIOREDOXIN-LIKE PROTEIN FROM BRADYRHIZOBIUM-JAPONICUM, ALTERS ITS REDOX PROPERTIES BUT NOT ITS IN-VIVO FUNCTIONS

TlpA, the membrane-anchored, thioredoxin-like protein from Bradyrhizob ium japonicum, is essential for cytochrome aa(3) biogenesis, The perip lasmic domain of TlpA was previously shown to have protein thiol:disul fide oxidoreductase activity and reducing properties similar to those of cytoplasmic thioredoxins, Here, we replaced the proline-109 in its active-site sequence (CVPC110)-V-107-P-108-C-109 by a histidine residu e, The resulting active-site motif (CVHC) resembles that of oxidizing thiol:disulfide oxidoreductases such as protein disulfide isomerase (P DI) and DsbA. Indeed, the TlpA variant P109H was by 66 mV more oxidizi ng than the wild-type protein, Nevertheless, the altered protein was e ven more efficient in catalyzing the reduction of insulin disulfides b y dithiothreitol than the wild-type due to a faster recycling of its c atalytically active, reduced form, Cells of B. japonicum expressing on ly the mutated tlpA gene had the same phenotypes as mild-type cells, s uggesting that the change in the redox potential of TlpA was not criti cal for its in vivo function. (C) 1997 Federation of European Biochemi cal Societies.