Specialization of a calmodulin-like protein: Androcam adopts a single conformation over the entire physiological range of calcium
The ubiquitous and highly conserved calcium binding protein calmodulin exhibits structural plasticity, broad target binding specificity and the ability to tune its affinity for Ca2+ ion. Collectively, these properties enable calmodulin to transduce biological calcium signals to hundreds of downstream targets. Despite the versatility of calmodulin, metazoans express many essential calmodulin-like proteins that perform tissue specific functions. In this thesis, I have studied androcam, an essential protein in D.melanogaster that is 67% identical to calmodulin, to determine how its structure, Ca+2 binding and target recognition properties differ from those of calmodulin and contribute to its unique function. My NMR structures solved at high and low calcium show that unlike calmodulin, which switches its conformation in response to changes in [Ca+2], each lobe of androcam is locked in a single fold over the entire physiological range of [Ca+2]. The androcam C-lobe has two EF hands which each ligand a Ca+2 ion, and is structurally similar to calmodulin. However, it binds Ca+2 much tighter than calmodulin and is therefore constitutively present in the Ca+2 bound "open" conformation that potentiates interactions with hydrophobic targets. The N lobe of androcam does not bind Ca+2 at physiological concentrations but is well structured and adopts a "closed" conformation similar to Ca+2-free calmodulin that is not expected to bind a hydrophobic anchor. Consistent with these structural observations, chemical shift perturbation experiments show that androcam interacts with the unique 'Insert2' peptide of the biological target Myosin VI with its C lobe only, whereas calmodulin binds this target using both lobes. Our results indicate that the androcam sequence has been optimized by evolution starting from the highly versatile calmodulin sequence to adopt only one of the many conformations that calmodulin can sample. We propose that many other calmodulin-like proteins might have also evolved to be specialists for a unique functional state out of the plethora of conformations that calmodulin has been shown to populate.
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