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Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF

In this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C‐terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N‐terminal domain generated a mutant protein with more than twofold increased...

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Detalles Bibliográficos
Autores principales: Fang, Zhen, Zhang, Juan, Liu, Baihong, Du, Guocheng, Chen, Jian
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4720410/
https://www.ncbi.nlm.nih.gov/pubmed/26552936
http://dx.doi.org/10.1111/1751-7915.12300
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author Fang, Zhen
Zhang, Juan
Liu, Baihong
Du, Guocheng
Chen, Jian
author_facet Fang, Zhen
Zhang, Juan
Liu, Baihong
Du, Guocheng
Chen, Jian
author_sort Fang, Zhen
collection PubMed
description In this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C‐terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N‐terminal domain generated a mutant protein with more than twofold increased catalytic activity towards casein. Replacement of the C‐terminal domain obviously improved keratinolytic activity and increased the k(cat)/K(m) value on a synthetic peptide, succinyl‐Ala‐Ala‐Pro‐Phe‐p‐nitroanilide, by 54.5%. Replacement of both the N‐ and C‐terminal domains generated a more stable mutant protein, with a T (m) value of 64.60 ± 0.65°C and a half‐life of 244.6 ± 2 min at 60°C, while deletion of the C‐terminal domain from KerSMD or KerSMF resulted in mutant proteins exhibiting high activity under mesophilic conditions. These findings indicate that the pre‐peptidase C‐terminal domain and N‐propeptide are not only important for substrate specificity, correct folding and thermostability but also support the ability of the enzyme to convert feather waste into feed additives.
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spelling pubmed-47204102016-01-29 Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF Fang, Zhen Zhang, Juan Liu, Baihong Du, Guocheng Chen, Jian Microb Biotechnol Research Articles In this study, we enhanced the catalytic efficiency and thermostability of keratinase KerSMD by replacing its N/C‐terminal domains with those from a homologous protease, KerSMF, to degrade feather waste. Replacement of the N‐terminal domain generated a mutant protein with more than twofold increased catalytic activity towards casein. Replacement of the C‐terminal domain obviously improved keratinolytic activity and increased the k(cat)/K(m) value on a synthetic peptide, succinyl‐Ala‐Ala‐Pro‐Phe‐p‐nitroanilide, by 54.5%. Replacement of both the N‐ and C‐terminal domains generated a more stable mutant protein, with a T (m) value of 64.60 ± 0.65°C and a half‐life of 244.6 ± 2 min at 60°C, while deletion of the C‐terminal domain from KerSMD or KerSMF resulted in mutant proteins exhibiting high activity under mesophilic conditions. These findings indicate that the pre‐peptidase C‐terminal domain and N‐propeptide are not only important for substrate specificity, correct folding and thermostability but also support the ability of the enzyme to convert feather waste into feed additives. John Wiley and Sons Inc. 2015-11-10 /pmc/articles/PMC4720410/ /pubmed/26552936 http://dx.doi.org/10.1111/1751-7915.12300 Text en © 2015 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Articles
Fang, Zhen
Zhang, Juan
Liu, Baihong
Du, Guocheng
Chen, Jian
Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title_full Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title_fullStr Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title_full_unstemmed Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title_short Enhancement of the catalytic efficiency and thermostability of S tenotrophomonas sp. keratinase KerSMD by domain exchange with KerSMF
title_sort enhancement of the catalytic efficiency and thermostability of s tenotrophomonas sp. keratinase kersmd by domain exchange with kersmf
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4720410/
https://www.ncbi.nlm.nih.gov/pubmed/26552936
http://dx.doi.org/10.1111/1751-7915.12300
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