Antimicrobial Peptide Variants and Polynucleotides Encoding Same

Abstract

The present invention relates to variants of a parent antimicrobial peptide. The present invention also relates to polynucleotides encoding the variants; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and methods of using the variants.

What is claimed is: 
     
         1 . An isolated variant of an antimicrobial peptide having the amino acid sequence of SEQ ID NO: 2, comprising a substitution at one or more positions 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 15, 17, 19, and 21 of the amino acid sequence of SEQ ID NO: 2; wherein the variant has antimicrobial activity, and the number of substitutions is 1-11, wherein when said variant comprises a substitution selected from G1R or G1K, said variant further comprises a substitution at positions 5 or 17. 
     
     
         2 . The variant of  claim 1 , wherein the number of substitutions are 1-10, 1-9 substitutions, 1-8 substitutions, 1-7 substitutions, 1-6 substitutions, 1-5 substitutions, 1-4 substitutions, 1-3 substitutions, or 1-2 substitutions. 
     
     
         3 . The variant of any  claim 1 , which further comprises one or more (several) substitutions selected from the group consisting of
 G1A, G1D, G1F, G1H, G1I, G1K, G1M, G1Q, G1R, G1S, G1T, G1V, G1W, G1Y,   F2A, F2G, F2H, F2I, F2L, F2M, F2P, F2S, F2V, F2W, F2Y,   C3L,   W4A, W4E, W4F, W4G, W41, W4L, W4M, W4N, W4Q, W4S, W4T, W4V, W4Y,   Y5E, Y5F, Y5G, Y5H, Y5K, Y5N, Y5R, Y5S, Y5W,   V6A, V6C, V6E, V6F, V6G, V6H, V61, V6L, V6M, V6N, V6Q, V6R, V6S, V6T, V6W, V6Y, C7V,   V8A, V8F, V8G, V8H, V81, V8L, V8N, V85, V8T, V8W, V8Y,   Y9A, Y9D, Y9F, Y9G, Y9H, Y91, Y9K, Y9M, Y9Q, Y9R, Y9S, Y9T, Y9V, Y9W,   R10K, R10P, R10S, R10T,   N11A, N11G, N11H, N11Q, N11R, N115, N11Y,   G12A, G12D, G12E, G12F, G12H, G12K, G12N, G12R, G125, G12Y,   V13A, V13C, V13F, V13G, V13H, V13K, V13L, V13P, V13Q, V13R, V135, V13T, V13W, V13Y,   V15A, V15C, V15F, V15G, V15H, V15I, V15K, V15L, V15M, V15N, V15P, V15Q, V15R, V155,   V15T, V15W, V15Y,   Y17C, Y17F, Y17G, Y17H, Y17I, Y17K, Y17L, Y17M, Y17N, Y17Q, Y17R, Y17S, Y17T, Y17V,   Y17W,   R19D, R19H, R19K, R19M, R19T, R19Y,   N21A, N21C, N21F, N21G, N21H, N21I, N21K, N21L, N21M, N21P, N21Q, N21R, N21S,   N21T, N21W, and N21Y.   
     
     
         4 . The variant of  claim 1 , which comprises a substitution at two of positions 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 15, 17, 19, and 21. 
     
     
         5 . The variant of  claim 1 , which comprises a substitution at three of positions 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 15, 17, 19, and 21. 
     
     
         6 . The variant of  claim 1 , which comprises a substitution at four of positions 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 15, 17, 19, and 21. 
     
     
         7 . The variant of  claim 1 , which comprises a substitution at one or more (several) of positions
 1, 2, 4, 5, 6, 8, 9, 12, 13, 15, 17, and 21.   
     
     
         8 . The variant of  claim 1 , which comprises the amino acid sequence of any of SEQ ID NO: 3-9, 13-25, 29-46, 50-55, 57, 59-60, 62, 65-66, 79-85, 88-91, 93-94, 96-116, 118-119, 126, 128-129, 138-160, 167, 178, 183, 196-197, 205, 212-214, 216-219, 338-344, 347-349, 352-353, 356-357, 361-362, 369-370, 383, 390, 392, 494, 500, and 543. 
     
     
         9 . The variant of  claim 1 , which comprises the amino acid sequence of any of SEQ ID NO: 13-25, 46, 57, 59-60, 66, 89, 94, 96-116, 118, 129, 138-160, 167, 178, 183, 196-197, 205, 212-214, 216-219, 338-344, 347-349, 352-353, 356-357, 361-362, 369-370, 383, 390, 392, 494, 500, and 543. 
     
     
         10 . An isolated polynucleotide encoding the variant of  claim 1 . 
     
     
         11 . A nucleic acid construct comprising the polynucleotide of  claim 10 . 
     
     
         12 . An expression vector comprising the polynucleotide of  claim 10 . 
     
     
         13 . A host cell comprising the polynucleotide of  claim 10 . 
     
     
         14 . A method of producing a variant of an antimicrobial peptide having the amino acid sequence of SEQ ID NO: 2, comprising:
 a) cultivating the host cell of  claim 13  under conditions suitable for the expression of the variant; and   b) recovering the variant.   
     
     
         15 . A method of treating a microbial infection in a subject in need thereof comprising administering an amount of the variant of  claim 1  effective to treat said microbial infection. 
     
     
         16 . A composition comprising the variant of  claim 1  and an antibiotic.