PURIFICATION AND CHARACTERIZATION OF ELASTASE FROM LEUKOCYTES CELLS  ISOLATED FROM MASTITIS MILK

S. Doosh, K.; M. A. Salih, A.; A. Shaker, K.

	PURIFICATION AND CHARACTERIZATION OF ELASTASE FROM LEUKOCYTES CELLS ISOLATED FROM MASTITIS MILK
IRAQ JOURNAL OF AGRICULTURE
Article 1, Volume 14, Issue 8, 2009, Pages 0-0
Authors
K. S. Doosh; A. M. A. Salih; K. A. Shaker
Abstract
Elastase was isolated from polymorphonuclear leukocytes and purified by the Ion-exchange column Diethyl amino ethyl Cellulose (DEAE-Cellulose), followed by gel filtration on Sephadex G-100 column, purification folds and the enzyme yield was 8.06 and 18.15% respectively. The burity of obtained enzyme was tested by polyacrylamide gel electrophoresis with sodium dodecyle sulfate and indicated that it has been purified to homogeneity by giving a single band. The results of enzyme characterization showed that the molecular weights were 22500 and 23000 Daltons as determined by gel filtration and electrophoresis in the presence of SDS .The optimum pH for the enzyme activity was 8.0 and was stable at pH values ranged between 7.0-9.0. The enzyme exhibited the maximum activity at 40ºC. The study of heat stability pointed out that the enzyme retained entire activity over 30 min. incubation at 30 - 50 Cº and remained active even when incubated at 20-80ºC. It was noticed that the addition of each of CaCl2, HgCl2 didn’t affect enzyme activity while CuCl2, NaCL reduced enzyme activity by 50 and 10 % respectively. However the other reagent which were specific inhibitors for cysteine, aspartic, metallo protease didn’t affect enzymo activity, mean while the enzyme completely inhibited by PMSF (specific serin protease inhibitor) so enzyme could be identify as serine proteases (Elastase).

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