The elusive role of the N-terminal extension of βA3- and βAl-crystallin

β-Crystallins are structural lens proteins with a conserved two-domain structure and variable N- and C-terminal extensions. These extensions are assumed to be involved in quaternary interactions within the β-crystallin oligomers or with other lens proteins. Therefore, the production of βA3- and βAl-crystallin from the single βA3/A1 mRNA by dual translation initiation is of interest. These crystallins are identical, except that βAl has a much shorter N-terminal extension than βA3. This rare mechanism has been conserved for over 250 million years during the evolution of the βA3/A1 gene, suggesting that the generation of different N-terminal extensions confers a selective advantage. We therefore compared the stability and association behaviour of recombinant βA3- and βAl-crystallin. Both proteins are equally stable in urea- and pH-induced denaturation experiments. Gel filtration and analytical ultracentrifugation established that βA3 and βA1 both form homodimers. In the water-soluble proteins of bovine lens, βA3 and βA1 are present in the same molecular weight fractions, indicating that they oligomerize equally with other β-crystallins. 1H-NMR spectroscopy showed that residues Met1 to Asn22 of the N-terminal extension of βA3 have great flexibility and are solvent exposed, excluding them from protein interactions in the homodimer. These results indicate that the different N-terminal extensions of βA3 and βA1 do not affect their homo- or heteromeric interactions.