Studies on the Active Site of Leukotriene A4 Hydrolase

Leukotriene (LT) A4 hydrolase (EC 3.3.2.6) is a soluble monomeric protein with a molecular mass of 69 kDa that converts the unstable epoxide intermediate LTA4 into the proinflammatory compound LTB4, 5(S), 12(R)-dihydroxy-6, 14-cis-8, 10-trans-eicosa-tetraenoic acid (For reviews see Refs. 1 and 2). Sequence comparisons of LTA4 hydrolase with certain zinc containing proteases and peptidases led to the discovery of a zinc binding motif in the primary structure of the enzyme3. Further studies verified that LTA4 hydrolase indeed contained one zinc atom per enzyme molecule4,5. These findings also led to the discovery of a previously unknown peptidase/amidase activity5,6 which was specifically stimulated by monovalent anions, e. g., chloride ions7, and also by albumin8. Studies involving site directed mutagenesis in combination with metal determination of the purified mutated proteins demonstrated that the zinc atom was bound to His-295, His-299, and Glu-318, in accordance with previous predictions, and that replacements of either of these three residues led to enzymatically inactive proteins which did not contain zinc9.