engleski

Yeast dipeptidyl peptidase III – insight into the structure of metallopeptidases of M49 family

Dipeptidyl peptidases III (DPP III) are zinc-dependent enzymes that cleave the first two amino acids from N terminus of different length peptides. The DPP III (M49) family is a relatively new metallopeptidase family (MEROPS: the protease database, http://www.merops.ac.uk), whose members are grouped on the basis of their sequence relationships and the unique HEXXGH motif at the active site. The exact physiological roles of DPPs III are still unclear and may be complex. Besides implied role of DPP III in intracellular protein catabolism, involvement in the pain-modulation and in endogenous defense against oxidative stress is indicated. In order to further characterise DPP III protein, biochemical, kinetic and structural properties of the yeast ortholog were investigated. The gene YOL057W encoding the DPP III from yeast Saccharomyces cerevisiae was expressed in homologous and heterologous system, along with several mutant forms. Stable and high yield expression of DPP III monomer (C130S mutant) enabled crystallization and determination of the 3D structure. The high-resolution crystal structure (1.95 Å) of DPP III apoenzyme reveals a novel protein fold, with two domains separated by a cleft, representing the prototype for the M49 family of peptidases. Although the overall structure of DPP III is completely unrelated, the structure of the active site is rather similar with that of thermolysin and neprilysin, two other zinc peptidases. Based on this and modeled structure of yeast DPP III in complex with poly-Ala octapeptide, the catalytic mechanism and mode of substrate binding was proposed. High degree of conservation especially in the active site cleft between yeast and mammalian DPP III allows correlation of structural, biochemical and physiological data for further investigation of these metallopeptidases.

yeast dipeptidyl peptidase III; crystal structure; M49 family

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