Isolation and molecular cloning of carbohydrate binding module (CBM40) from Vibrio cholerae Non-O1 Neuraminidase

Shadariah M., . and Nadiawati A., . Isolation and molecular cloning of carbohydrate binding module (CBM40) from Vibrio cholerae Non-O1 Neuraminidase. pp. 108-118. ISSN 2180-1983

Carbohydrate binding modules (CBMs) are discrete contiguous amino acid sequence within a carbohydrate-active enzyme which are non-catalytic modules that primarily exist to target parent enzyme to its substrate for efficient hydrolysis. Although many sialidase proteins have been identified from various pathogenic bacteria only a few enzymes are commercially available which have been used for chemoenzymatic syntheses and therapeutics application. In order to study family 40 CBM domain a number of bacteria have been screened including Pseudomonas aeruginosa ATCC 27853 Bacillus cereus ATCC 14579 Staphylococcus aureus ATCC 33862 Salmonella thypii ATCC 14028 and Vibrio cholerae Non-O1. A gene encoding CBM40 domain was screened from all the bacteria strains and subjected to PCR amplification. From all the samples tested only Vibrio cholerae Non-O1 amplified a PCR product with approximate size of 530 bp. From BLAST sequence analysis result has shown 99 similarity with the target Vibrio cholerae neuraminidase NanH (M83562). Next the confirmed CBM40 gene was further ligated into pGEMT Easy Vector system and transformed into E. coli JM109 host to secure the clone before religated into suitable expression vector.

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Abstract

Carbohydrate binding modules (CBMs) are discrete contiguous amino acid sequence within a carbohydrate-active enzyme which are non-catalytic modules that primarily exist to target parent enzyme to its substrate for efficient hydrolysis. Although many sialidase proteins have been identified from various pathogenic bacteria only a few enzymes are commercially available which have been used for chemoenzymatic syntheses and therapeutics application. In order to study family 40 CBM domain a number of bacteria have been screened including Pseudomonas aeruginosa ATCC 27853 Bacillus cereus ATCC 14579 Staphylococcus aureus ATCC 33862 Salmonella thypii ATCC 14028 and Vibrio cholerae Non-O1. A gene encoding CBM40 domain was screened from all the bacteria strains and subjected to PCR amplification. From all the samples tested only Vibrio cholerae Non-O1 amplified a PCR product with approximate size of 530 bp. From BLAST sequence analysis result has shown 99 similarity with the target Vibrio cholerae neuraminidase NanH (M83562). Next the confirmed CBM40 gene was further ligated into pGEMT Easy Vector system and transformed into E. coli JM109 host to secure the clone before religated into suitable expression vector.

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Item Type:	Article
AGROVOC Term:	Carbohydrates
AGROVOC Term:	Amino acid sequences
AGROVOC Term:	Isolation
AGROVOC Term:	Molecular cloning
AGROVOC Term:	Extraction
AGROVOC Term:	Electrophoresis
AGROVOC Term:	Enzymatic hydrolysis
AGROVOC Term:	PCR
AGROVOC Term:	Polymerase chain reaction
AGROVOC Term:	Purification
Depositing User:	Ms. Suzila Mohamad Kasim
Last Modified:	24 Apr 2025 06:29
URI:	http://webagris.upm.edu.my/id/eprint/24975

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