PMID: 2113042Mar 1, 1990Paper

Semisynthesis of carboxy-terminal fragments of thermolysin

International Journal of Peptide and Protein Research
V De Filippis, A Fontana

Abstract

Enzyme-catalyzed synthesis of two polypeptide fragments, one of which is obtained by chemical synthesis, in the presence of proteolytic enzymes and in aqueous organic solvents constitutes a convenient procedure for the synthesis of proteins and their analogs. This novel semisynthetic procedure was investigated for preparing COOH-terminal fragments of the metallo-protease thermolysin. Fragment 205-316, obtained by autolysis of the protein in the presence of EDTA, was first cleaved selectively with Staphylococcus aureus V8 protease at the level of the single Glu302 residue into fragments 205-302 and 303-316. Upon incubation for 2-5 days of fragment 205-302 with a 5-fold excess of peptide 303-316, prepared by solid phase synthesis, with V8-protease in 0.1 M ammonium acetate, pH 6.0, containing 50% glycerol as organic cosolvent, enzyme-catalyzed reformation of the peptide bond was achieved in yields up to approximately 90% (based on fragment 205-302). The same procedure was used to prepare also the thermolysin fragments 205-315 and 205-311 by enzymatic coupling of fragment 205-302 to peptide 303-315 or 303-311, these last prepared by proteolytic digestion of the synthetic peptide 303-316. This procedure of semisynthesis opens up an...Continue Reading

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Citations

May 4, 2000·Protein Science : a Publication of the Protein Society·S KumaranR P Roy
May 22, 2002·Protein Science : a Publication of the Protein Society·Sonati Srinivasulu, A Seetharama Acharya
Feb 15, 1992·European Journal of Biochemistry·I Svendsen, K Breddam
Jan 1, 1994·Peptides·J Bongers, E P Heimer
Aug 1, 1991·Current Opinion in Biotechnology·J HumphriesR A Smith

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