E. coli methionine sulfoxide reductase with a truncated N terminus or C terminus, or both, retains the ability to reduce methionine sulfoxide.

The monomeric peptide methionine sulfoxide reductase (MsrA) catalyzes the irreversible thioredoxin-dependent reduction of methionine sulfoxide. The crystal structure of MsrAs from Escherichia coli and Bos taurus can be described as a central core of about 140 amino acids that contains the active site. The core is wrapped by two long N- and C-terminal extended chains. The catalytic mechanism of the E. coli enzyme has been recently postulated to take place through formation of a sulfenic acid intermediate, followed by reduction of the intermediate via intrathiol-disulfide exchanges and thioredoxin oxidation. In the present work, truncated MsrAs at the N- or C-terminal end or at both were produced as folded entities. All forms are able to reduce methionine sulfoxide in the presence of dithiothreitol. However, only the N-terminal truncated form, which possesses the two cysteines located at the C-terminus, reduces the sulfenic acid intermediate in a thioredoxin-dependent manner. The wild type displays a ping-pong mechanism with either thioredoxin or dithiothreitol as reductant. Kinetic saturation is only observed with thioredoxin with a low K(M) value of 10 microM. Thus, thioredoxin is likely the reductant in vivo. Truncations do not significantly modify the kinetic properties, except for the double truncated form, which displays a 17-fold decrease in k(cat)/K(MetSO). Alternative mechanisms for sulfenic acid reduction are also presented based on analysis of available MsrA sequences.