IIB   20738
INSTITUTO DE INVESTIGACIONES BIOLOGICAS
Unidad Ejecutora - UE
artículos
Título:
Peptide synthesis catalyzed by a haloalkaliphilic serine protease from the archaeon Natrialba magadii
Autor/es:
RUIZ, D. M.; IANNUCI, N.; CASCONE, O.; DE CASTRO, R.E.
Revista:
LETTERS IN APPLIED MICROBIOLOGY
Editorial:
WILEY-BLACKWELL PUBLISHING, INC
Referencias:
Año: 2010 vol. 51 p. 691 - 696
ISSN:
0266-8254
Resumen:
Aims: Haloarchaeal proteases function optimally in high salt (low water activity); thus, they offer an advantage over the nonhalophilic counterparts as biocatalysts for protease-catalysed peptide synthesis. The haloalkaliphilic archaeonHaloarchaeal proteases function optimally in high salt (low water activity); thus, they offer an advantage over the nonhalophilic counterparts as biocatalysts for protease-catalysed peptide synthesis. The haloalkaliphilic archaeon Natrialba magadii secretes a solvent-tolerant protease, Nep (Natrialba magadiisecretes a solvent-tolerant protease, Nep (Natrialba magadii extracellular protease). In this work, the ability of Nep to catalyse peptide synthesis was examined. Methods and Results: The tripeptide Ac-Phe-Gly-Phe-NH2 was synthesized using Ac-Phe-OEt and Gly-Phe-NH2 substrates as building blocks in the presence of Nep, 30% (v ⁄ v) dimethyl sulfoxide (DMSO) and 1Æ5 or 0Æ5 mol l)1The tripeptide Ac-Phe-Gly-Phe-NH2 was synthesized using Ac-Phe-OEt and Gly-Phe-NH2 substrates as building blocks in the presence of Nep, 30% (v ⁄ v) dimethyl sulfoxide (DMSO) and 1Æ5 or 0Æ5 mol l)12 substrates as building blocks in the presence of Nep, 30% (v ⁄ v) dimethyl sulfoxide (DMSO) and 1Æ5 or 0Æ5 mol l)1⁄ v) dimethyl sulfoxide (DMSO) and 1Æ5 or 0Æ5 mol l)1 NaCl. Purification and identification of the peptide product was achieved by RP-HPLC and ESI-MS, respectively. The native as well as the recombinant enzyme produced in Haloferax volcanii (HvNep) was similarly effective as catalysts for the synthesis of this model tripeptide with yields of up to 60% and without secondary hydrolysis of the product. HvNep catalysed the synthesis of various tripeptides with preference for those having aromatic amino acids in the P1 site.Haloferax volcanii (HvNep) was similarly effective as catalysts for the synthesis of this model tripeptide with yields of up to 60% and without secondary hydrolysis of the product. HvNep catalysed the synthesis of various tripeptides with preference for those having aromatic amino acids in the P1 site.HvNep catalysed the synthesis of various tripeptides with preference for those having aromatic amino acids in the P1 site.