Journal of Stress Physiology &
Biochemistry, Vol. 9 No. 1
2013, pp. 184-208 ISSN 1997-0838
Original Text Copyright (cc) 2013 by Kapoor, Tripathi and Shrivastava
ORIGINAL
ARTICLE
Data source : Google Scholar
QueryDate : 2016-12-24
Cites : 2
Isolation and Purification of
Heterotetrameric Catalase from a Desiccation Tolerant Cyanobacterium Lyngbya arboricola
Shivali Kapoor* , S. N. Tripathi,
Alpana Shrivastava
Department of Botany, Banaras Hindu University, Varanasi 221005,
India
*E-Mail: shivalihm@gmail.com
Received October 25, 2012
The desiccation
tolerant cyanobacterium Lyngbya arboricola, isolated from bark surfaces
of Mangifera indica,
possessed up to four stable isoforms of catalase in addition to other
antioxidative enzymes, for several years under a dry state.
Purification of the two most persistent isoforms of catalase (Cat) has
been undertaken by employing acetone precipitation, ethanol: chloroform
treatment, gel filtration and ion exchange chromatography. The two
isoforms of catalase remained almost unchanged on varying matric and
osmotic hydration levels of mats of the cyanobacterium. The
purification procedures resulted in a 1.3 % yield of purified single
isoform (0.22 mg mL-1 protein) with 709 Units mg-1
specific activity and a purity index of 0.83. Five millimolar of
dithiothreitol (DTT) was observed to be pertinent in maintaining the
optimum redox state of the enzyme. The purification procedures
additionally facilitated the simultaneous elimination and procurement
of phycoerythrins (PE) and mycosporine-like amino acids (MAA). Each
purified isoform gave a single band (~45kDa) upon SDS-PAGE and
denaturing urea isoelectric focusing (IEF) depicted the presence of 2
subunits each of CatA and CatB. The monoisotopic mass and pI value of
CatA and CatB as revealed by LC-MS analysis and internal amino acid
sequencing was 78.96, 5.89 and 80.77, 5.92, respectively, showing
resemblance with CatA of Erysiphe
graminis subs. hordei
and CatB of Ajellomyces
capsulata. The heterotetrameric monofunctional catalase (~320
kDa), due to its stability in the form of resistance to ethanol:
chloroform, its thermoalkaliphilic nature and the presence of
innumerable hydrophobic amino acid residues (~40%), thus exhibited its
potential for biotechnological applications.
