Semisynthesis of human growth hormone‐releasing factor by trypsin catalyzed coupling of leucine amide to a C‐terminal acid precursor

Abstract

Human growth hormone‐releasing factor, GRF(1–44)‐NH₂, was synthesized by trypsin catalyzed coupling of Leu‐NH₂ to Arg⁴³ of the precursor, GRF(1–43)‐OH, prepared by solid phase peptide synthesis. The semisynthetic GRF(1–44)‐NH₂ was fully characterized and showed full potency in the rat pituitary in vitro bioassay. Conversion to GRF(1–44)‐NH₂ was limited to 60–70% in both 75% v:v N,N′‐dimethylacetamide and 95% v:v 1,4‐butanediol due to competing transpeptidations at Arg⁴¹ and Arg³⁸ generating [Leu⁴²]‐GRF(1–42)‐NH₂ and [Leu³⁹]‐GRF(1–39)–NH₂ side‐products, respectively. The rates of formation and yields of GRF(1–44)‐NH₂ versus pH, Leu‐NH₂ concentration, and solvent composition were also studied.