Crystal Structure And Function Of The Phosphorylation Of Threonine Lyase Spvc

MAPKs are evolutionarily conserved and play vital roles in both animal and plantinnate immune responses. Therefore one of the common strategies used by pathogenicbacteria is to interfere with MAPK signaling to evade host immune defenses.Recent studies indicate that, SpvC, conserved in both animal and plant pathogenicbacteria, belongs to a novel family of enzymes designated phosphothreonine lyase thatirreversibly inactivate mitogen-activated protein kinases (MAPKs) to repress host innateimmunity. In contrast with protein phosphatases, phosphothreonine lyase recognizes thephosphothreonine in pThr-Xaa-pTyr motif (Xaa stands for any residue) contained withinthe activated MAPKs and removes its phosphate moiety by cleaving the C-OP rather thanthe O-P bond, thus permanently inactivating them. Although phosphothreonine lyases donot exhibit activity toward phosphotyrosine in pThr-Xaa-pTyr motif, they do favor thebiphosphorylated form over the monophosphorylated form. Substitution ofphosphothreonine in pThr-Xaa-pTyr motif with phosphoserine compromises both peptidesubstrate recognition and catalytic efficiency of OspF, indicating that phosphothreoninelyase displays a preference for phosphothreonine over phosphoserine to remove thephosphate group.To study the reaction mechanism catalyzed by this family of enzymes, we determinedthe crystal structures of SpvC. The crystal structure of SpvC reveals a novel protein foldcontaining a single compactα/βdomain. The core of SpvC is composed of sevenanti-parallelβ-strands and eightα-helices. A shallow but prominent groove on the surfaceof SpvC formed by loops L1, L2 and L3, and the twisted 5-stranded is likely thesubstrate-binding site. Two adjacent basic surfaces formed by conserved residues among SpvC family enzymes may play an important role in recognizing the phosphate groups inthe biphosphorylated substrate as further evidenced by the mutagenesis assay. Theinvariant K136 in SpvC is proposed to be the catalytic residue responsible for theelimination reaction. These results not only provide significant insights into the reactionmechanism catalyzed by this family of enzymes, but also suggest a potential drug bindingpocket that can be used to design small molecular inhibitor for therapeutic agents.