Single nucleotide polymorphisms (SNPs) are present in the global transcriptional regulator cyclic AMP (cAMP) receptor protein (CRP) of the attenuated vaccine strain Mycobacterium bovis, bacillus CalmetteGuérin (BCG). We have found that these SNPs resulted in small but significant changes in the expression of a number of genes in M. tuberculosis when a deletion of the Rv3676 CRP was complemented by the BCG allele, compared to complementation by the M. tuberculosis allele. We can explain these changes in gene expression by modeling the structure of the mycobacterial protein on the known structure of CRP from Escherichia coli. Thus, the SNP change in the DNA-binding domain, Lys178, is predicted to form a hydrogen bond with the phosphate backbone of the DNA, as does the equivalent residue in E. coli, whereas Glu178 in M. tuberculosis/M. bovis does not, thus explaining the stronger binding reported for CRP of BCG to CRP-binding sites in mycobacterial DNA. In contrast, the SNP change in the nucleotide binding domain (Leu47Pro) is predicted to result in the loss of one hydrogen bond, which is accommodated by the structure, and would not therefore be expected to cause any change in function relating to cAMP binding. The BCG allele fully complemented the growth defect caused by the deletion of the Rv3676 protein in M. tuberculosis, both in vitro and in macrophage and mouse infections, suggesting that these SNPs do not play any role in the attenuation of BCG. However, they may have allowed BCG to grow better under the in vitro-selective conditions used in its derivation from the M. bovis wild type.Tuberculosis (TB), caused by the bacterium Mycobacterium tuberculosis, remains the infectious disease that kills the largest number of people worldwide, despite the fact that a vaccine, bacillus Calmette-Guérin (BCG), has been available since the 1920s and is relatively inexpensive. BCG is the world's most widely used vaccine, but it does not provide complete protection against TB. At most, it provides only 80% protection (14), and is particularly effective against disseminated forms of TB in children, such as miliary TB and tuberculosis meningitis. However, the protection conferred against pulmonary TB in adults, the majority of carriers of the disease burden, has been very variable in clinical trials, depending on the population, the country, and the BCG strain used. BCG does not in fact provide adequate levels of protection in Africa, India, and some parts of the United States. So, new vaccines are required, and a great deal of current research is directed toward achieving this end (4), including the use of BCG as the starting point for new vaccines (20). It is therefore important to have as much information as possible about the genomic and virulence status of the BCG strain to know how it differs from virulent strains.Using the attenuated strain vaccine production techniques pioneered by Louis Pasteur, BCG was derived by Calmette and Guérin from virulent Mycobacterium bovis, the agent of bovine TB and a close relative of M....