Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4

Most fuel cells nowadays use platinum as the electrocatalyst, but this metal is expensive and difficult to be obtained. This gives rise to many studies on enzymatic fuel cells. The major hurdle in enzymatic fuel cell is that the hydrogen oxidation at anode is sensitive to oxygen, and this requires a...

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Main Author:	Hussin, Mohd. Adhhaa
Format:	Thesis
Published:	2014
Subjects:	QD Chemistry
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spelling	my-utm-ep.483972017-08-23T04:54:52Z Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4 2014 Hussin, Mohd. Adhhaa QD Chemistry Most fuel cells nowadays use platinum as the electrocatalyst, but this metal is expensive and difficult to be obtained. This gives rise to many studies on enzymatic fuel cells. The major hurdle in enzymatic fuel cell is that the hydrogen oxidation at anode is sensitive to oxygen, and this requires an oxygen-insensitive enzyme as the catalyst. [NiFe] hydrogenase, comprised of two subunits, is the best known hydrogenase to be used as biocatalyst since it is insensitive to oxygen. [NiFe] hydrogenase genes are organized in an operon, containing a cluster of genes needed for the maturation of the enzyme. This study was initiated to detect and amplify the genes in putative [NiFe] hydrogenase operon in Anoxybacillus sp. SK3-4 by PCR and DNA sequencing. The sequences of the amplified genes were comfirmed by alignment with the published sequences in the NCBI database. 11 pairs of primer were designed for both PCR and sequencing, based on the full genome sequence of Anoxybacillus sp. SK3-4. All the primers designed are 18-22 nucleotides in length with a minimum GC content of 50%. The difference in Tm between the primers pairs are in the range of 0.1-2.3oC. It can be comfirmed that the putative [NiFe] hydrogenase operon consists of 11 genes that are transcribed into one polycistronic mRNA with 10 101 nucleotides and translated into 11 different proteins. Futher sequence analysis showed that [NiFe] hydrogenase in Anoxybacillus sp. SK3-4 is predicted to be classified into group 1 hydrogenase, which is a membrane-bound hydrogenase. The large subunit protein is the most important protein in the [NiFe] hydrogenase operon where the catalytic activity resdes. The large subunit has 57% similarity with [NiFe] hydrogenase from Allochromatum Vinosum (PDB ID 3MYR). A homology model has been constructed for Anoxybacillus sp. SK3-4 [NiFe] hydrogenase using the template structure of A. vinosum. The large subunit consist of 1720 nucleotides that codes for 573 amino acids with a molecular weight of 64.7 kDa. The theoritical pI has been predicted to be 6.25. 2014 Thesis http://eprints.utm.my/id/eprint/48397/ masters Universiti Teknologi Malaysia, Faculty of Bioscience and Medical Engineering Faculty of Bioscience and Medical Engineering
institution	Universiti Teknologi Malaysia
collection	UTM Institutional Repository
topic	QD Chemistry
spellingShingle	QD Chemistry Hussin, Mohd. Adhhaa Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
description	Most fuel cells nowadays use platinum as the electrocatalyst, but this metal is expensive and difficult to be obtained. This gives rise to many studies on enzymatic fuel cells. The major hurdle in enzymatic fuel cell is that the hydrogen oxidation at anode is sensitive to oxygen, and this requires an oxygen-insensitive enzyme as the catalyst. [NiFe] hydrogenase, comprised of two subunits, is the best known hydrogenase to be used as biocatalyst since it is insensitive to oxygen. [NiFe] hydrogenase genes are organized in an operon, containing a cluster of genes needed for the maturation of the enzyme. This study was initiated to detect and amplify the genes in putative [NiFe] hydrogenase operon in Anoxybacillus sp. SK3-4 by PCR and DNA sequencing. The sequences of the amplified genes were comfirmed by alignment with the published sequences in the NCBI database. 11 pairs of primer were designed for both PCR and sequencing, based on the full genome sequence of Anoxybacillus sp. SK3-4. All the primers designed are 18-22 nucleotides in length with a minimum GC content of 50%. The difference in Tm between the primers pairs are in the range of 0.1-2.3oC. It can be comfirmed that the putative [NiFe] hydrogenase operon consists of 11 genes that are transcribed into one polycistronic mRNA with 10 101 nucleotides and translated into 11 different proteins. Futher sequence analysis showed that [NiFe] hydrogenase in Anoxybacillus sp. SK3-4 is predicted to be classified into group 1 hydrogenase, which is a membrane-bound hydrogenase. The large subunit protein is the most important protein in the [NiFe] hydrogenase operon where the catalytic activity resdes. The large subunit has 57% similarity with [NiFe] hydrogenase from Allochromatum Vinosum (PDB ID 3MYR). A homology model has been constructed for Anoxybacillus sp. SK3-4 [NiFe] hydrogenase using the template structure of A. vinosum. The large subunit consist of 1720 nucleotides that codes for 573 amino acids with a molecular weight of 64.7 kDa. The theoritical pI has been predicted to be 6.25.
format	Thesis
qualification_level	Master's degree
author	Hussin, Mohd. Adhhaa
author_facet	Hussin, Mohd. Adhhaa
author_sort	Hussin, Mohd. Adhhaa
title	Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
title_short	Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
title_full	Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
title_fullStr	Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
title_full_unstemmed	Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4
title_sort	detection and amplification of putative [nife] hydrogenase operon in anoxybacillus sp. sk3-4
granting_institution	Universiti Teknologi Malaysia, Faculty of Bioscience and Medical Engineering
granting_department	Faculty of Bioscience and Medical Engineering
publishDate	2014
_version_	1747817380309893120

Detection and amplification of putative [NiFe] hydrogenase operon in anoxybacillus sp. SK3-4

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