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Keratinases from pseudomonas aeruginosa KS-1: Characterization and degradation of surrogate prion protein Sup 35NM

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Title Keratinases from pseudomonas aeruginosa KS-1: Characterization and degradation of surrogate prion protein Sup 35NM
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Contributor Gupta, Rani
 
Subject Microbiology
surrogate
protein
 
Description Keratinases are a special group of proteases which are well recognized for their action
newlineon recalcitrant proteins such as feather, nail, hair, hoof etc. They are lately being recognized
newlineas potential catalysts for degradation of prion. Few keratinases have already been
newlinedocumented to degrade prion however treatment of the infected tissue with harsh alkali or at
newlinehigh temperature is required. This limits the applicability of these enzymes. Thus, the need of
newlinethe hour is to search for efficient keratinases which can degrade prion under ambient
newlineconditions.
newlineLooking into the above, the present investigation was undertaken to search for a
newlinepotential prion degrading keratinase. Fifty feather degrading bacterial isolates were screened,
newlineout of which eleven were selected as they could degrade chicken feather within 48 h. Among
newlinethese, extracellular broth of nine isolates carried out cell free feather degradation. Isolate KS-
newline1 which maximally degraded Sup 35NM was finally selected for further studies. It was
newlineidentified as Pseudomonas aeruginosa and deposited to MTCC under the accession number
newline10775.
newlineTwo keratinases, KP1 and KP2 of 45kDa and 33kDa respectively were purified from
newlineits extracellular broth. Keratinase KP1 shared homology with putative aminopeptidase while
newlinekeratinase KP2 gave homology with pseudolysin. Keratinase KP1 was alkaline, serine, metal
newlineactivated protease and KP2 was a neutral, thiol-activated, serine protease. Both the enzymes
newlinewere extremely thermostable. K:C (keratinolytic: caseinolytic) ratio of 2.5 and 0.9 for KP1
newlineand KP2 respectively. Both also efficiently hydrolyzed p-nitroanilides with best activity on
newlineN-Suc-Ala-Ala-Pro-Phe-pNA. They had unique activity on the synthetic substrate of plasmin.
newlineHydrolysis of insulin B chain revealed KP1 and KP2 to have four and six cleavage sites
newlinerespectively.
newlineBoth the keratinases were constitutively expressed as extracellular proteins using
newlinepEZZ18-E. coli HB101 system. Biochemical characterization the pH and temperature optima
newlineof rKP1 and rKP2 to be pH 10.0/60oC
Bibliography p.196-207
 
Date 2014-04-28T12:01:03Z
2014-04-28T12:01:03Z
2014-04-28
n.d.
2012
n.d.
 
Type Ph.D.
 
Identifier http://hdl.handle.net/10603/18010
 
Language English US
 
Relation -
 
Rights university
 
Format 207p.
-
None
 
Coverage Microbiology
 
Publisher New Delhi
University of Delhi
Dept. of Microbiology
 
Source INFLIBNET