Synthesis of the segment (11–23) located in the first tandem repeat of plasma kallikrein: comparative binding studies of this and another segment (328–343) to high-molecular-mass kininogen

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Abstract

The synthesis of porcine plasma kallikrein (pPK) segment (11–23), of sequence Phe-Phe-Arg-Gly-Gly-Asp-Val-Ser-Ala-Met-Tyr-Thr-Pro, present in the first tandem repeat sequence of the regulatory chain of PK, has been accomplished following the peptide fragments (5 + 4 + 4) condensation strategy in solution, as well as by fluorenylmethoxycarbonyl solid-phase chemistry. This and another synthetic PK segment of residues (328–343) present in the fourth tandem repeat sequence [Cys(ACM)-Ser-Leu-Arg-Leu-Ser-Thr-Asp-Gly-Ser-Pro-Thr-Arg-Ile-Thr-Tyr] and synthesized by a solid-phase method, were fully characterized by 1H nuclear magnetic resonance, fast atom bombardment mass spectrometry, amino acid composition and reversed-phase high-performance liquid chromatography. Proteolysis of these peptides by either rat PK (rPK) or trypsin resulted in cleavages between Arg↓Gly for pPK (11–23) and between Arg↓Leu and Arg↓Ile for rPK (328–343). Kinetic studies revealed that for peptide pPK (11–23), the catalytic efficiency (kcat/Km) of rPK is ≅ 9-fold higher than that of trypsin, but for the other peptide, rPK (328–343), kcat/Km of trypsin is ≅ 49-fold higher than that of rPK. The facile cleavage of pPK (11–23) by rPK confirms the Arg13↓Gly14 position as the site of autolytic degradation of PK and also explains its special preference for Phe-Phe-Arg sequence.

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    This paper was presented in part at the 33rd Annual Meeting of the Canadian Federation of Biological Societies, Halifax, June 1990, Abstract No. 224, p. 73.

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