Investigation of the effect of freezing on protease-catalyzed peptide synthesis using cryoprotectants and frozen organic solvent |
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Authors: | Haensler M Arnold K |
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Institution: | Institute of Biochemistry, Faculty of Biosciences, Pharmacy and Psychology, University of Leipzig, Germany. |
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Abstract: | In order to investigate the effect of freezing on aqueous protease-catalyzed peptide synthesis systems, the influence of polyethylene glycols as cryoprotecting substances on alpha-chymotrypsin-catalyzed coupling of a N-protected acyl donor ester and various nucleophilic amino components was studied. Changes in S'-specificity of alpha-chymotrypsin in frozen aqueous systems were suppressed by polyethylene glycols even at concentrations below 1% (w/v). Furthermore, the influence of freeze-concentration in organic solvents on protease-catalyzed peptide synthesis was investigated for the first time. In frozen tert-butanol, alpha-chymotrypsin-catalyzed peptide synthesis took advantage from freeze-concentration, but in contrast to frozen aqueous systems, no changes in S'-specificity of the biocatalyst were observed. The results suggest that freeze-concentration is not the only cause of freezing-induced yield improvement in aqueous peptide synthesis systems, but interactions between enzyme and ice structures strongly contribute to the observed effects. |
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