Amino acid residues that are critical for in vivo catalytic activity of CtpA, the carboxyl-terminal processing protease for the D1 protein of photosystem II

CtpA, a carboxyl-terminal processing protease, is a member of a novel famil y of endoproteases that includes a tail-specific protease from Escherichia coli. In oxygenic photosynthetic organisms, CtpA catalyzes C-terminal proce ssing of the D1 protein of photosystem II, an essential event for the assem bly of a manganese cluster and consequent light-mediated water oxidation. W e introduced site-specific mutations at 14 conserved residues of CtpA in th e cyanobacterium Synechocystis sp. PCC 6803 to examine their functional rol es. Analysis of the photoautotrophic growth capabilities of these mutants, their ability to process precursor D1 protein and hence evolve oxygen, alon g with an estimation of the protease content in the mutants revealed that f ive of these residues are critical for in vivo activity of CtpA. Recent x-r ay crystal structure analysis of CtpA from the eukaryotic alga Scenedesmus obliquus (Liao, D.-I., Qian, J., Chisholm, D. A., Jordan, D. B. and Diner, B. A. (2000) Nat. Struct. Biol. 7, 749-753) has shown that the residues equ ivalent to Ser-313 and Lys-338, two of the five residues mentioned above, f orm the catalytic center of this enzyme. Our in vivo analysis demonstrates that the three other residues, Asp-253, Arg-255, and Glu-316, are also impo rtant determinants of the catalytic activity of CtpA.