Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5

Genome sequence of Indonesian probiotic of Lactobacillus plantarum II1A5 contains a gene encoding a proteolytic subunit of caseinolytic protease, designated as ClpP_LP. This study aims to express the Clp gene heterological and apply its proteolytic activity to some livestock products. To address th...

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Autores principales:	M. Yusuf, C. Budiman, I. I. Arief, C. Sumantri
Formato:	article
Lenguaje:	EN
Publicado:	IPB University 2021
Materias:	Clp-Protease heterologous expression Lactobacillus plantarum IIA-1A5 structural homology modelling synthetic gene Animal culture SF1-1100
Acceso en línea:	https://doaj.org/article/79dfb66d3d7943bd9d6ad16be03895a4
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spelling	oai:doaj.org-article:79dfb66d3d7943bd9d6ad16be03895a42021-11-15T23:17:05ZSynthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A510.5398/tasj.2021.44.4.5202615-787X2615-790Xhttps://doaj.org/article/79dfb66d3d7943bd9d6ad16be03895a42021-11-01T00:00:00Zhttps://journal.ipb.ac.id/index.php/tasj/article/view/34024https://doaj.org/toc/2615-787Xhttps://doaj.org/toc/2615-790X Genome sequence of Indonesian probiotic of Lactobacillus plantarum II1A5 contains a gene encoding a proteolytic subunit of caseinolytic protease, designated as ClpP_LP. This study aims to express the Clp gene heterological and apply its proteolytic activity to some livestock products. To address this, the gene encoding ClpP_LP was optimized in silico by improving its Codon Adaptation Index and GC content to 0.94 and 53.62%, respectively. The optimized gene was then inserted into pET28a, transformed into Escherichia coli BL21(DE3), and over-expressed by induction of 1 mM Isopropyl β-D-1-thiogalactopyranoside at 37°C. The result showed that ClpP_LP was successfully over-expressed in a fully soluble form with the specific activity towards milk casein was 7739.89 AU mg-1. This activity was significantly greater than that of chymotrypsin. Further, the three-dimensional model of ClpP_LP was built using SWISS MODEL, which showed that this protein formed a homo-tetradecameric (14-mer) structure with each monomer consisting of 7 α-helix and 10 β-sheets. The identification of the active side showed that the active side of ClpP_LP is Ser-97, His-122, Asp-171, and forms a substrate-binding cavity with a size of about 29.5 Ǻ. Overall, our approach can serve as an appropriate platform for the production of ClpP_LP in a large-scale production for various applications in dairy products and derivatives. M. YusufC. BudimanI. I. AriefC. SumantriIPB UniversityarticleClp-Proteaseheterologous expressionLactobacillus plantarum IIA-1A5structural homology modellingsynthetic geneAnimal cultureSF1-1100ENTropical Animal Science Journal, Vol 44, Iss 4 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Clp-Protease heterologous expression Lactobacillus plantarum IIA-1A5 structural homology modelling synthetic gene Animal culture SF1-1100
spellingShingle	Clp-Protease heterologous expression Lactobacillus plantarum IIA-1A5 structural homology modelling synthetic gene Animal culture SF1-1100 M. Yusuf C. Budiman I. I. Arief C. Sumantri Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
description	Genome sequence of Indonesian probiotic of Lactobacillus plantarum II1A5 contains a gene encoding a proteolytic subunit of caseinolytic protease, designated as ClpP_LP. This study aims to express the Clp gene heterological and apply its proteolytic activity to some livestock products. To address this, the gene encoding ClpP_LP was optimized in silico by improving its Codon Adaptation Index and GC content to 0.94 and 53.62%, respectively. The optimized gene was then inserted into pET28a, transformed into Escherichia coli BL21(DE3), and over-expressed by induction of 1 mM Isopropyl β-D-1-thiogalactopyranoside at 37°C. The result showed that ClpP_LP was successfully over-expressed in a fully soluble form with the specific activity towards milk casein was 7739.89 AU mg-1. This activity was significantly greater than that of chymotrypsin. Further, the three-dimensional model of ClpP_LP was built using SWISS MODEL, which showed that this protein formed a homo-tetradecameric (14-mer) structure with each monomer consisting of 7 α-helix and 10 β-sheets. The identification of the active side showed that the active side of ClpP_LP is Ser-97, His-122, Asp-171, and forms a substrate-binding cavity with a size of about 29.5 Ǻ. Overall, our approach can serve as an appropriate platform for the production of ClpP_LP in a large-scale production for various applications in dairy products and derivatives.
format	article
author	M. Yusuf C. Budiman I. I. Arief C. Sumantri
author_facet	M. Yusuf C. Budiman I. I. Arief C. Sumantri
author_sort	M. Yusuf
title	Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
title_short	Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
title_full	Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
title_fullStr	Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
title_full_unstemmed	Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5
title_sort	synthetic gene-based heterologous expression, proteolytic, and structural characterization of caseinolytic protease of lactobacillus plantarum iia-1a5
publisher	IPB University
publishDate	2021
url	https://doaj.org/article/79dfb66d3d7943bd9d6ad16be03895a4
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Synthetic Gene-Based Heterologous Expression, Proteolytic, and Structural Characterization of Caseinolytic Protease of Lactobacillus plantarum IIA-1A5

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