PROLYL 4-HYDROXYLASES AND THEIR PROTEIN DISULFIDE-ISOMERASE SUBUNIT

Prolyl 4-hydroxylases (EC 1.14.11.2) catalyze the formation of 4-hydro xyproline in collagens and other proteins with collagen-like sequences . The vertebrate type I and type II enzymes are [alpha(I)](2) beta(2) and [alpha(II)](2) beta(2) tetramers, respectively, whereas the enzyme from the nematode Caenorhabditis elegans is an alpha beta dimer. The type I enzyme is the major form in most but not all vertebrate tissues . The catalytic properties of the various enzyme forms are highly simi lar, but there are distinct, although small, differences in K-m values for various peptide substrates between the enzyme forms and major dif ferences in K-i values for the competitive inhibitor, poly(L-proline). Prolyl 4-hydroxylase requires Fe2+, 2-oxoglutarate, O-2 and ascorbate . Kinetic studies and theoretical considerations have led to elucidati on of the reaction mechanism, and recent extensive site-directed mutag enesis studies have identified five critical residues at the cosubstra te binding sites. A number of compounds have been characterized that i nhibit it competitively with respect to some of the cosubstrates, and three groups of suicide inactivators have also been identified. The be ta subunit in all forms of prolyl 4-hydroxylase is identical to protei n disulfide isomerase (PDI), a multifunctional polypeptide that also s erves as a subunit in the microsomal triglyceride transfer protein, as a chaperone-like polypeptide that probably assists folding of a numbe r of newly synthesized proteins, and in several other functions. The m ain role of the PDI polypeptide as a protein subunit is probably relat ed to its chaperone function. Recent expression studies of recombinant human prolyl di-hydroxylase subunits in a yeast have indicated that t he formation of a stable enzyme tetramer in vivo requires coexpression of collagen polypeptide chains.