A conserved noncanonical motif in the pseudoactive site of the ROP5 pseudokinase domain mediates its effect on Toxoplasma virulence

J Biol Chem. 2011 Jun 27. [Epub ahead of print]

A conserved noncanonical motif in the pseudoactive site of the ROP5 pseudokinase domain mediates its effect on Toxoplasma virulence.

Reese ML, Boothroyd JC.

SourceStanford University, United States.

Abstract
The ROP5 family is a closely related set of polymorphic pseudokinases that are critical to Toxoplasma's ability to cause disease. ROP5's polymorphisms also make it a major determinant of strain-specific differences in virulence. ROP5 possesses all of the major kinase motifs required for catalysis except for a substitution at the catalytic Asp. We show that this substitution in ROP5's catalytic loop is part of a motif conserved in other pseudokinases of both Toxoplasma and human origin, and that this motif is required for ROP5's full activity in vivo. This suggests evolutionary selection at this site for a biochemical function, rather than simple drift away from catalysis. We present the crystal structures of a virulent isoform of ROP5 both in its ATP-bound and -unbound states and have demonstrated that, in spite of maintaining the canonical ATP-binding motifs, ROP5 binds ATP in a distorted conformation mediated by unusual Mg coordination sites that would not be predicted from the primary sequence. In addition, we have mapped the polymorphisms spread throughout ROP5's primary sequence to two major surfaces, including ROP5's activation segment. This suggests that the pseudoactive site of this class of pseudokinases may have evolved to use the canonical ATP-binding motifs for non-catalytic signaling through allostery.

PMID:21708941[PubMed - as supplied by publisher]