Molecular cloning, structure and bait region splice variants of α₂-macroglobulin from the soft tick Ornithodoros moubata

The sequence of a α₂-macroglobulin (α₂M) from the soft tick Ornithodoros moubata (TAM) was determined by cloning and sequencing of overlapping polymerase chain reaction (PCR) and rapid amplification of cDNA ends PCR products. The TAM cDNA sequence is 4944 bp long and contains one open reading frame coding for a protein precursor composed of 1494 amino-acid residues, including a 24-residue signal sequence. The mature protein is cleaved into two subunits similarly to the C3 and C4 components of complement and fish α₂Ms. Phylogeny analysis revealed that TAM is closely related to Limulus α₂M and displays the highest similarity to the partial sequence of α₂M from hard tick Ixodes scapularis. The comparison of conserved cysteine residues between TAM and human and Limulus α₂Ms made it possible to predict the pattern of disulfide bridges and explain the atypical molecular arrangement of TAM. Four variants of the TAM bait region differing only in a short central segment were found; our data indicate that TAM exists as a single-copy gene in the tick genome and its bait region variants likely arise by alternative splicing. TAM is produced by tick hemocytes and it is also significantly expressed in salivary glands. TAM mRNA levels were shown to be up-regulated upon blood meal.