Functional chimeras between the catalytic domains of the mycobacterial adenylyl cyclase Rv1625c and a Paramecium guanylyl cyclase

Abstract

The class IIIa adenylyl cyclase (AC) Rv1625c from Mycobacterium tuberculosis forms homodimers with two catalytic centres, whereas the Paramecium guanylyl and mammalian ACs operate as pseudoheterodimers with one catalytic centre. The functional and structural relationship of the catalytic domains of these related class III cyclases was investigated. Point mutations introduced into Rv1625c to engineer a forskolinbinding pocket created a single heterodimeric catalytic centre, yet did not result in forskolin activation. Chimerization of these Rv1625c point mutants with corresponding mammalian AC domains was impossible. However, it was successful using a complemental Paramecium guanylyl cyclase domain and resulted in an AC. The data signify a divergence of structural and functional evolution in class III Acs.

Mycobacterial adenylyl cyclases: Biochemical diversity and structural plasticity

“…Interestingly, the C2‐like Rv1625c mutant protein (D300A) could heterodimerize and reconstitute activity with the Paramaecium guanylyl cyclase C1‐like domain (the second cyclase domain in this guanylyl cyclase, but functionally a C1 domain) [20]. However, Rv1625c mutants could not reconstitute activity with C1 or C2 domains of the mammalian adenylyl cyclase isoforms [20], or with Rv1647 as described below.…”

A Structural Basis for the Role of Nucleotide Specifying Residues in Regulating the Oligomerization of the Rv1625c Adenylyl Cyclase from M.tuberculosis

“…The diverse nature of the adenylyl cyclases from M. tuberculosis is apparent from the unique biochemical properties displayed by purified proteins in vitro. [13][14][15][16] Subtle structural plasticity could be the source of the biochemical divergence between enzymes from the same organism. Understanding these structural differences would also provide an opportunity to design rational drugs that target only the bacterial enzymes without affecting its eukaryotic counterparts.…”

Interaction of Rv1625c, a mycobacterial class IIIa adenylyl cyclase, with a mammalian congener

“…This indicates that the mycobacterial and mammalian AC membrane domains either do not share a common ancestor or that they have degenerated beyond recognition. Although many biochemical properties of the catalytic homodimer of AC Rv1625c resemble those of mammalian ACs (Guo et al ., 2001), the generation of an active cyclase dimer from a mammalian and an AC Rv1625c catalytic domain is not possible (Linder et al ., 2004a). This indicates incompatible structural differences which preclude a productive interaction of the mycobacterial and mammalian catalytic domains.…”