Stabilization of T4 endonuclease V

Abstract

The invention provides T4 endonuclease V compositions that exhibit enhanced stability, including stability at non-refrigerated temperatures, through reduced activity of cryptic proteases. Methods for detecting cryptic protease activity and methods for reducing such activity are also provided.

1 . A composition comprising T4 endonuclease V wherein: 
 (a) the composition has T4 endonuclease V activity; and    (b) a sample of the composition when tested for protease activity using a labeled reporter peptide whose amino acid sequence is Pro Leu Ser Arg Thr Leu Ser Val Ala Ala Lys (SEQ ID NO:1) gives a percentage of degraded reporter peptide of less than 10 percent, where the test for protease activity comprises the steps of:    (i) preparing a test solution comprising an aqueous buffer, a sample of the composition, and the labeled reporter peptide, where the concentration of the T4 endonuclease V in the test solution is 0.095±0.05 mg/ml;    (ii) incubating the test solution at 25° C. for a period of 7 days; and    (iii) determining the percentage of degraded reporter peptide after step (ii) by subjecting at least a portion of the test solution to a peptide separation procedure and determining the percentage of degraded reporter peptide by densitometry.    
     
     
         2 . The composition of  claim 1  wherein a sample of the composition when tested for storage stability at 25° C. for a test period of four weeks retains over that period at least 10 percent of its T4 endonuclease V activity at the beginning of the period.  
     
     
         3 . The composition of  claim 1  wherein a sample of the composition when tested for storage stability at 25° C. for a test period of six months retains over that period at least 10 percent of its T4 endonuclease V activity at the beginning of the period.  
     
     
         4 . A composition comprising T4 endonuclease V wherein: 
 (a) the composition has T4 endonuclease V activity; and    (b) the composition shows no difference by statistical analysis (p>0.05) in specific T4 endonuclease V activity between storage for one week at 4° C. and storage for one week at 25° C.