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Original
Research Article
Production
of Extracellular Anti-leukaemic Enzyme L-asparaginase
from Marine Actinomycetes by Solid-state and Submerged
Fermentation: Purification and Characterisation
N Saleem
Basha1*, R Rekha2, M Komala1
and S Ruby3
1Department
of Pharmaceutical Biotechnology, 2Department
of Pharmacognosy, 3Department of
Pharmaceutical Chemistry, Mohamed Sathak A J College of
Pharmacy, Medavakkam, Chennai 600 119, India.
*Corresponding
author: E-mail: nsaleem_basha@rediffmail.com
Tel:
+91449943998065 (mobile);
Fax:
+ 91-44-24502573
Received: 23 November
Revised
accepted: 16 May 2009
Tropical
Journal of Pharmaceutical Research, August 2009; 8(4):
353-360
Abstract
Purpose:
The objective of this investigation was to isolate
marine actinomycetes, screen them for L-asparaginase
activity and characterise the enzyme.
Methods:
Marine actinomycetes were isolated from sediment samples
obtained from Tamilnadu and Kerala in India. The
isolates were identified as actinomycetes by
microscopical and biochemical tests. Production of L-asparaginase
was carried out in three different media, namely,
solid-state media, Tryptone Glucose Yeast extract (TGY)
broth and Tryptone Fructose Yeast extract (TFY) broth..
The enzyme was purified to near homogeneity by ammonium
sulphate precipitation, dialysis, gel filtration on
Sephadex G-100 column and SDS-PAGE.
Results:
Among 10 marine isolates subjected to preliminary
screening, only isolates S3, S4 and K8 showed potential
for L-asparaginase activity. All three marine soil
isolates synthesized asparaginase with yield ranging
from 24.6 to 49.2 IU/ml. Soil isolate S3 showed the
highest productivity of 49.2 IU/ml with a protein
content of 65 µg/ml and optimum activity at pH 7.5 and
50 ºC. The apparent Km value for the
substrate was 25µM. Mg2+ ion slightly
stimulated activity while Cu2+, Zn2+
and EDTA were inhibitory.
Conclusion:
The study revealed that marine actinomycetes may be a
potential source of high yield, high substrate
specificity L-asparaginase, which is an anti-leukaemia
agent.
Key words:
L-asparaginase, Solid-state media, TGY broth, TFY broth,
SDS-PAGE, Km and Vmax. |
