Identification of proteins in the exosporium of Bacillus anthracis

Abstract

Spores of Bacillus anthracis, the causative agent of anthrax, possess an exosporium. As the outer surface layer of these mature spores, the exosporium represents the primary contact surface between the spore and environment/host and is a site of spore antigens. The exosporium was isolated from the endospores of the B. anthracis wild-type Ames strain, from a derivative of the Ames strain cured of plasmid pXO2 " , and from a previously isolated pXO1" doubly cured strain, B. anthracis UM23Cl2. The protein profiles of SDS-PAGE-separated exosporium extracts were similar for all three. This suggests that avirulent variants lacking either or both plasmids are realistic models for studying the exosporium from spores of B. anthracis. A number of loosely adsorbed proteins were identified from amino acid sequences determined by either nanospray-MS/MS or N-terminal sequencing. Salt and detergent washing of the exosporium fragments removed these and revealed proteins that are likely to represent structural/integral exosporium proteins. Seven proteins were identified in washed exosporium: alanine racemase, inosine hydrolase, ExsF, CotY, ExsY, CotB and a novel protein, named ExsK. CotY, ExsY and CotB are homologues of Bacillus subtilis outer spore coat proteins, but ExsF and ExsK are specific to B. anthracis and other members of the Bacillus cereus group.

The role of a purine-specific nucleoside hydrolase in spore germination of Bacillus thuringiensis

“…The amino acid sequence of BC2889 is almost identical to the amino acid sequence of the putative NH purified from exosporia of B. cereus and B. anthracis (Redmond et al, 2004;Steichen et al, 2003;Todd et al, 2003). Based on the sequence of BC2889 and its homologues found in members of the B. cereus group, a 1.4 kb DNA fragment was amplified from B. thuringiensis genomic DNA by PCR.…”

“…NHs are crucial in the salvage pathway of protozoan parasites which are deficient in the pathway for de novo synthesis of purine nucleotides (Gopaul et al, 1996). Non-parasitic NHs, for example the salivary purine nucleosidase of the mosquito Aedes aegypti, may be involved in the degradation of modified nucleosides found in tRNA (Giabbai & Degano, 2004;Ribeiro & Valenzuela, 2003), but apart from this, no clear role for non-parasitic NHs, including the NH identified from the exosporia of Bacillus anthracis and Bacillus cereus, has been reported so far (Giabbai & Degano, 2004;Redmond et al, 2004;Ribeiro & Valenzuela, 2003;Todd et al, 2003).…”

“…The outermost layer of the B. cereus group spore is a loose-fitting, balloon-like structure known as an exosporium, which consists mainly of protein, polysaccharides, lipids and ash (Matz et al, 1970). Dozens of proteins, including a putative NH, were identified from the exosporia of B. cereus and B. anthracis spores (Redmond et al, 2004;Steichen et al, 2003;Todd et al, 2003). Although the role of some proteins has been studied, most functions remain to be elucidated (Boydston et al, 2006;Ramarao & Lereclus, 2005;Steichen et al, 2005;Yan et al, 2007).…”

Lab-Scale production of Bacillus atrophaeus' spores by solid state fermentation in fifferent types of bioreactors

“…Knowing that the growth rate of the microorganism depends on the local environmental conditions (Redmond et al, 2004;Vries et al, 2005), the evaluation of spore's yields from Erlenmeyer flasks and plastic bags fermentation was done by comparative kinetic studies. During fermentation kinetic in Erlenmeyer flasks the pH of the culture had an increased 1.6 in seven days incubation and decrease 0.5 from the 7th to the 9th incubation day, the water activity of the substrate (0.97) did not change, the amount of moisture content (wet-basis) has remained at 87%, the difference between the calculated moisture (91%) and the obtained (87%) is due to loss of water during sterilization process.…”

Analysis of a novel spore antigen in Bacillus anthracis that contributes to spore opsonization

“…This antibody was provided to the Critical Reagent Program by the Identification Group, Detection Department, Defence Science and Technology Laboratory (Dstl), Porton Down, Wiltshire, UK. This antibody, BA-MAB5, was developed against unfixed purified exosporia (Redmond et al, 2004) from the B. anthracis Ames strain. While the precise epitope of BA-MAB5 has not been mapped, it recognizes a protein of~250 kDa and is presumed to be BclA.…”