Carboxypeptidase A Loss Of Crystallographic Studies Of The Living Mechanism

The structures of the complexes of Carboxypeptidase A (CPA) with inactivator have been determined by X-ray crystallography. The inactivator, 2-benzyl-3-iodopropanoic acid binds covalently to CPA(Glu-270) at the 2 complexes. The 2-benzyl-3-iodopropanoic acid complexes with CPA crystallize in the space group P21 and P212121, with unit cell parameters a=74.5, b=60.5, c = 65.6, β=98.1° for P21 and a=48.8, b= 66.9, c=96.0 for P212121. A total of 34092 unique reflections were collected to 2.0A|° resolution on Mac-Sience DIP-2000 area detector for P21 and 10767 unique reflections to 2.5 A resolution for P212121. Both structures were determined by molecular replacement with AMoRe of the CPP4 proramm package using the CPA from the natural crystal as the search model due to the large cell parameters difference between natural CPA and the complexes. The final crystallographic residuals are 0.163 for P21 and 0.152 for P2₁2₁2₁. The inactivators exhibits normal binding mode compared with other complexes of CPA: the benzyl occupies hydrophobic pocket; the carboxylate of inactivator forms a salt link with the positively charged guanidinium moiety of Arg145 and a hydrogen bond donated from Tyr-248 and a hydrogen bond donated from Asn144. But surprisingly, the distance between carboxylate of inactivator and Zn bound water is within the range of hydrogen bond, which was not seen in the other complexes of CPA.In the complexes, the torsion angle C2-C3-Glu270OE2-GluCD is 175° ,179°in P21 complexes and -174° in P2₁2₁2₁, and the torsion angle GluO270E2-C3-C2-C2A is 92°, 78°for P21 and -47° for P2₁2₁2₁ respectively, which probably mean the attacking direction of Glu270 is different. We think it is likely the 2 complexes undergo different reaction mechanism in the details beacause of the difference of...