Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell

Glucosamine (GlcN) is a widely used food supplement. Hence, enormous attention has been concerned with enzymatic production of GlcN owing to its advantage over a chemical approach. In this study, a previously unstudied chitinase gene (MxChi) in the genome of <i>Myxococcus xanthus</i> was...

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Autores principales:	Yongmei Lyu, Feng Zheng, Chuanxing Qiu, Meng Wang, Dujun Wang, Xiaoyang Zhang, Josef Voglmeir, Li Liu, Xiaohong Yu
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	glucosamine chitinase <i>M. xanthus</i> chitin enzymatic cascades Chemical technology TP1-1185
Acceso en línea:	https://doaj.org/article/c86b135d1f694c768717bbcf9bb5c2c1
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spelling	oai:doaj.org-article:c86b135d1f694c768717bbcf9bb5c2c12021-11-25T17:35:52ZHeterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell10.3390/foods101128082304-8158https://doaj.org/article/c86b135d1f694c768717bbcf9bb5c2c12021-11-01T00:00:00Zhttps://www.mdpi.com/2304-8158/10/11/2808https://doaj.org/toc/2304-8158Glucosamine (GlcN) is a widely used food supplement. Hence, enormous attention has been concerned with enzymatic production of GlcN owing to its advantage over a chemical approach. In this study, a previously unstudied chitinase gene (MxChi) in the genome of <i>Myxococcus xanthus</i> was cloned, expressed in recombinant soluble form and purified to homogeneity. TLC-, UPLC-, and microplate-reader- based activity tests confirmed MxChi hydrolyzes colloidal chitin to chitobiose as sole product. The optimal catalytic pH and temperature of MxChi was identified as 7.0 and 55 °C, respectively. MxChi exhibited 80% activity after 72 h incubation at 37 °C. The site-directed mutagenesis revealed that the amino acids D323A, D325A, and E327A of MxChi were in the DXDXE catalytic motif of GH18. When coupled with β-N-acetylhexosaminidase (SnHex) and deacetylase (CmCBDA), the enzyme allowed one-pot extraction of GlcN from colloidal chitin and shrimp shell. The optimal condition was 37 °C, pH 8.0, and 1/3/16.5 (MxChi/SnHex/CmCBDA), conducted by orthogonal design for the enzymatic cascades. Under this condition, the yield of GlcN was 26.33 mg from 400 mg shrimp shell. Facile recombinant in <i>E. coli</i>, robust thermostability and pure product herein makes newly discovered chitinase a valuable candidate for the green recycling of chitin rich waste.Yongmei LyuFeng ZhengChuanxing QiuMeng WangDujun WangXiaoyang ZhangJosef VoglmeirLi LiuXiaohong YuMDPI AGarticleglucosaminechitinase<i>M. xanthus</i>chitinenzymatic cascadesChemical technologyTP1-1185ENFoods, Vol 10, Iss 2808, p 2808 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	glucosamine chitinase <i>M. xanthus</i> chitin enzymatic cascades Chemical technology TP1-1185
spellingShingle	glucosamine chitinase <i>M. xanthus</i> chitin enzymatic cascades Chemical technology TP1-1185 Yongmei Lyu Feng Zheng Chuanxing Qiu Meng Wang Dujun Wang Xiaoyang Zhang Josef Voglmeir Li Liu Xiaohong Yu Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
description	Glucosamine (GlcN) is a widely used food supplement. Hence, enormous attention has been concerned with enzymatic production of GlcN owing to its advantage over a chemical approach. In this study, a previously unstudied chitinase gene (MxChi) in the genome of <i>Myxococcus xanthus</i> was cloned, expressed in recombinant soluble form and purified to homogeneity. TLC-, UPLC-, and microplate-reader- based activity tests confirmed MxChi hydrolyzes colloidal chitin to chitobiose as sole product. The optimal catalytic pH and temperature of MxChi was identified as 7.0 and 55 °C, respectively. MxChi exhibited 80% activity after 72 h incubation at 37 °C. The site-directed mutagenesis revealed that the amino acids D323A, D325A, and E327A of MxChi were in the DXDXE catalytic motif of GH18. When coupled with β-N-acetylhexosaminidase (SnHex) and deacetylase (CmCBDA), the enzyme allowed one-pot extraction of GlcN from colloidal chitin and shrimp shell. The optimal condition was 37 °C, pH 8.0, and 1/3/16.5 (MxChi/SnHex/CmCBDA), conducted by orthogonal design for the enzymatic cascades. Under this condition, the yield of GlcN was 26.33 mg from 400 mg shrimp shell. Facile recombinant in <i>E. coli</i>, robust thermostability and pure product herein makes newly discovered chitinase a valuable candidate for the green recycling of chitin rich waste.
format	article
author	Yongmei Lyu Feng Zheng Chuanxing Qiu Meng Wang Dujun Wang Xiaoyang Zhang Josef Voglmeir Li Liu Xiaohong Yu
author_facet	Yongmei Lyu Feng Zheng Chuanxing Qiu Meng Wang Dujun Wang Xiaoyang Zhang Josef Voglmeir Li Liu Xiaohong Yu
author_sort	Yongmei Lyu
title	Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
title_short	Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
title_full	Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
title_fullStr	Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
title_full_unstemmed	Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell
title_sort	heterologous expression of a thermostable chitinase from <i>myxococcus xanthus</i> and its application for high yield production of glucosamine from shrimp shell
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/c86b135d1f694c768717bbcf9bb5c2c1
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Heterologous Expression of a Thermostable Chitinase from <i>Myxococcus xanthus</i> and Its Application for High Yield Production of Glucosamine from Shrimp Shell

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