Title : Crystal structures of recombinant human
purple Acid phosphatase with and without an inhibitory conformation of the repression loop
Abstract :
- The crystal structure of human purple acid phosphatase recombinantly expressed in Escherichia coli ( rHPAP ( Ec )) and Pichia pastoris ( rHPAP ( Pp )) has been determined in two different crystal forms, both at 2.2A resolution
- In both cases, the enzyme crystallized in its oxidized (inactive) state, in which both Fe atoms in the dinuclear active site are Fe(III)
- The main difference between the two structures is the conformation of the enzyme "repression loop"
- Proteolytic cleavage of this loop in vivo or in vitro results in significant activation of the mammalian PAPs
- In the crystals obtained from rHPAP ( Ec ), the carboxylate side-chain of Asp145 of this loop acts as a bidentate ligand that bridges the two metal atoms, in a manner analogous to a possible binding mode for a phosphate ester substrate in the enzyme-substrate complex
- The carboxylate side-chain of Asp145 and the neighboring Phe146 side-chain thus block the active site , thereby inactivating the enzyme
- In the crystal structure of rHPAP ( Pp ), the enzyme "repression loop" has an open conformation similar to that observed in other mammalian PAP structures
- The present structures demonstrate that the repression loop exhibits significant conformational flexibility, and the observed alternate binding mode suggests a possible inhibitory role for this loop