Aims Examine the regulation of a spore coat protein and the effects on spore properties. Methods and Results A c. 23kDa band in coat/exosporial extracts of Bacillus anthracis Sterne spores varied in amount depending upon the conditions of sporulation. It was identified by MALDI as a likely orthologue of ExsB of Bacillus cereus. Little if any was present in an exosporial preparation with a location to the inner coat/cortex region established by spore fractionation and immunogold labelling of electron micrograph sections. Because of its predominant location in the inner coat, it has been renamed Cot gamma. It was relatively deficient in spores produced at 37 degrees C and when acidic fermentation products were produced a difference attributable to transcriptional regulation. The deficiency or absence of Cot gamma resulted in a less robust exosporium positioned more closely to the coat. These spores were less hydrophobic and germinated somewhat more rapidly. Hydrophobicity and appearance were rescued in the deletion strain by introduction of the cot gamma gene. Conclusions The deficiency or lack of a protein largely found in the inner coat altered spore hydrophobicity and surface appearance. Significance and Impact of the Study The regulated synthesis of Cot gamma may be a paradigm for other spore coat proteins with unknown functions that modulate spore properties in response to environmental conditions.