Prolyl 4-hydroxylases catalyze the formation of 4-hydroxyproline in collagens and other proteins with an appropriate collagen-like stretch of amino acid residues. The enzyme requires Fe(II), 2-oxoglutarate, molecular oxygen, and ascorbate. This review concentrates on recent progress toward understanding the detailed mechanism of 4-hydroxylase action, including: (a) occurrence and function of the enzyme in animals; (b) general molecular properties; (c) intracellular sites of hydroxylation; (d) peptide substrates and mechanistic roles of the cosubstrates; (e) insights into the development of antifibrotic drugs; (f) studies of the enzyme's subunits and their catalytic function; and (g) mutations that lead to Ehlers-Danlos Syndrome. An account of the regulation of collagen hydroxylase activities is also provided.