ABSTRACT
There continues to be a need for developing efficient and environmentally friendly treatments for
Bacillus anthracis
, the causative agent of anthrax. One emerging approach for inactivation of vegetative
B. anthracis
is the use of bacteriophage endolysins or lytic enzymes encoded by bacterial genomes (autolysins) with highly evolved specificity toward bacterium-specific peptidoglycan cell walls. In this work, we performed
in silico
analysis of the genome of
Bacillus anthracis
strain Ames, using a consensus binding domain amino acid sequence as a probe, and identified a novel lytic enzyme that we termed AmiBA2446. This enzyme exists as a homodimer, as determined by size exclusion studies. It possesses
N
-acetylmuramoyl-
l
-alanine amidase activity, as determined from liquid chromatography-mass spectrometry (LC-MS) analysis of muropeptides released due to the enzymatic digestion of peptidoglycan. Phylogenetic analysis suggested that AmiBA2446 was an autolysin of bacterial origin. We characterized the effects of enzyme concentration and phase of bacterial growth on bactericidal activity and observed close to a 5-log reduction in the viability of cells of
Bacillus cereus
4342, a surrogate for
B. anthracis
. We further tested the bactericidal activity of AmiBA2446 against various
Bacillus
species and demonstrated significant activity against
B. anthracis
and
B. cereus
strains. We also demonstrated activity against
B. anthracis
spores after pretreatment with germinants. AmiBA2446 enzyme was also stable in solution, retaining its activity after 4 months of storage at room temperature.
