Major surface protein 2 (MSP2) is an immunodominant and antigenically variant protein in the outer membrane of the rickettsia Anaplasma marginale. MSP2 variation is generated by recombination into a single operon-linked genomic expression site. The complete 5.6-kb msp2 locus was identified by sequencing a 90-kb region of the St. Maries strain of A. marginale. The locus encoded, in a 5' to 3' direction, a transcriptional regulator followed by five outer membrane proteins, OMP1, OpAG3, OpAG2, OpAG1, and MSP2. The sequences of this entire locus were analyzed using six genetically and phenotypically distinct strains of A. marginale. The overall locus structure was highly conserved with 100% identity among strains in the transcriptional regulator. Synonymous and nonsynonymous exchanges were infrequent in omp1 and rare in opag1 and opag2 among the six strains without strong bias for either type of exchange (neutral mutations). In contrast, mutations in opag3 seem to underlie purifying (negative) selection reflecting pressure to retain protein structure, in marked contrast to the highly antigenically variant MSP2. Interestingly, the 5' structure of this A. marginale msp2 locus is conserved in the omp1 gene locus of Ehrlichia chaffeensis and p30 gene locus of E. canis despite marked divergence between genera in the structure of the 3' region of the loci. This supports the hypothesis that the expression sites of these important immunogenic proteins are derived from a common precursor with later divergent evolution along genus lines.