Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation

Abstract. In the continual quest of novel biocatalysts with improved properties that will fulfil the raising industrial demands, directed evolution appears to be one of the most powerful tools in generating novel enzymes. Error -Prone PCR is a customized PCR protocol that will randomly introduce mut...

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Main Authors:	Ibrahim Ali, Noorbatcha, Hamzah, Mohd. Salleh
Format:	Conference or Workshop Item
Language:	English English English
Published:	2017
Subjects:	TP248.13 Biotechnology
Online Access:	http://irep.iium.edu.my/61791/ http://irep.iium.edu.my/61791/1/ARCSB%202017%20slides%20Bglk.pdf http://irep.iium.edu.my/61791/2/ARCSB%202017%20Timetable%209%20November%20%20%20%202017%20Bglk.pdf http://irep.iium.edu.my/61791/3/ARCSB2017%20-%20announcement.pdf

id	iium-61791
recordtype	eprints
spelling	iium-617912018-02-03T08:55:09Z http://irep.iium.edu.my/61791/ Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation Ibrahim Ali, Noorbatcha Hamzah, Mohd. Salleh TP248.13 Biotechnology Abstract. In the continual quest of novel biocatalysts with improved properties that will fulfil the raising industrial demands, directed evolution appears to be one of the most powerful tools in generating novel enzymes. Error -Prone PCR is a customized PCR protocol that will randomly introduce mutations creating a variant library. Beta glucoside kinase (bglk) is an enzyme that catalyzes the phosphor transformation to the 6’ C in a glucose molecule. The aim of the presented research is to engineer the bglk enzyme for phosphorus-sulfur bond formation. Two bglk libraries were created, one with a low mutation rate (0-4 mutation/kb) and the second with high mutation rate (9-16 mutation/kb) and was screened with Fluorescent Activated Cell Sorting (FACS) using a 6’thio-glucose-BODIPY as fluorescent substrate. Prior to this, a Glucose-BODIPY substrate was used with the wild type enzyme in order to test the cell incubation conditions with the fluorescent substrate. The positive clones collected from these two libraries, will be sequenced to verify the type and location of the mutations. 2017-11-22 Conference or Workshop Item PeerReviewed application/pdf en http://irep.iium.edu.my/61791/1/ARCSB%202017%20slides%20Bglk.pdf application/pdf en http://irep.iium.edu.my/61791/2/ARCSB%202017%20Timetable%209%20November%20%20%20%202017%20Bglk.pdf application/pdf en http://irep.iium.edu.my/61791/3/ARCSB2017%20-%20announcement.pdf Ibrahim Ali, Noorbatcha and Hamzah, Mohd. Salleh (2017) Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation. In: KUALA LUMPUR, Putrajaya Marriott Hotel, Malaysia. (Unpublished)
repository_type	Digital Repository
institution_category	Local University
institution	International Islamic University Malaysia
building	IIUM Repository
collection	Online Access
language	English English English
topic	TP248.13 Biotechnology
spellingShingle	TP248.13 Biotechnology Ibrahim Ali, Noorbatcha Hamzah, Mohd. Salleh Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
description	Abstract. In the continual quest of novel biocatalysts with improved properties that will fulfil the raising industrial demands, directed evolution appears to be one of the most powerful tools in generating novel enzymes. Error -Prone PCR is a customized PCR protocol that will randomly introduce mutations creating a variant library. Beta glucoside kinase (bglk) is an enzyme that catalyzes the phosphor transformation to the 6’ C in a glucose molecule. The aim of the presented research is to engineer the bglk enzyme for phosphorus-sulfur bond formation. Two bglk libraries were created, one with a low mutation rate (0-4 mutation/kb) and the second with high mutation rate (9-16 mutation/kb) and was screened with Fluorescent Activated Cell Sorting (FACS) using a 6’thio-glucose-BODIPY as fluorescent substrate. Prior to this, a Glucose-BODIPY substrate was used with the wild type enzyme in order to test the cell incubation conditions with the fluorescent substrate. The positive clones collected from these two libraries, will be sequenced to verify the type and location of the mutations.
format	Conference or Workshop Item
author	Ibrahim Ali, Noorbatcha Hamzah, Mohd. Salleh
author_facet	Ibrahim Ali, Noorbatcha Hamzah, Mohd. Salleh
author_sort	Ibrahim Ali, Noorbatcha
title	Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
title_short	Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
title_full	Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
title_fullStr	Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
title_full_unstemmed	Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation
title_sort	ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond formation
publishDate	2017
url	http://irep.iium.edu.my/61791/ http://irep.iium.edu.my/61791/1/ARCSB%202017%20slides%20Bglk.pdf http://irep.iium.edu.my/61791/2/ARCSB%202017%20Timetable%209%20November%20%20%20%202017%20Bglk.pdf http://irep.iium.edu.my/61791/3/ARCSB2017%20-%20announcement.pdf
first_indexed	2023-09-18T21:27:37Z
last_indexed	2023-09-18T21:27:37Z
_version_	1777412297705127936

Ultra-high-throughput screening of beta glucoside kinase random mutagenesis library for phosphorus-sulfur bond Formation

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