1.E.42 The Putative Holin-like Toxin (Hol-Tox) Family
The Putative Holin-like Toxin family consists of many small proteins (34-48aas) with a single TMSs. Rajesh et al. (2011) identified a holin-like gene from a goat skin surface metagenome and designated it tmp1. It coded for a 34 amino acyl peptide sharing sequence similarity with putative holin-like toxin genes. Following expression in E. coli, Tmp1 exhibited antibacterial activity against Gram-positive bacteria but not to Gram-negative bacteria. A single transmembrane domain (TMD) was identified within Tmp1, and deletion analysis of the N-terminal region and TMD indicated TMD to be responsible for the antibacterial activity. The TMD-dependent antibacterial activity was validated using a synthetic peptide with the amino acid sequence of TMD. Besides antibacterial activity, Tmp1 also complemented the function of holin in a lysis-defective bacteriophage lambda system. To broaden the spectrum of antibacterial activity, a mutant library of tmp1 was generated by random mutagenesis. Four mutants with amino acyl substitutions at the N-terminus of Tmp1 exhibited increased antibacterial activity against Gram-positive and Gram-negative bacteria but were not hemolytic. Improved activity of these mutant proteins was attributed to their increased hydrophobicity.