The binding of foot‐and‐mouth disease virus leader proteinase to eIF4GI involves conserved ionic interactions

Abstract

The leader proteinase (Lpro) of foot‐and‐mouth disease virus (FMDV) initially cleaves itself from the polyprotein. Subsequently, Lpro cleaves the host proteins eukaryotic initiation factor (eIF) 4GI and 4GII. This prevents protein synthesis from capped cellular mRNAs; the viral RNA is still translated, initiating from an internal ribosome entry site. Lpro cleaves eIF4GI between residues G674 and R675. We showed previously, however, that Lpro binds to residues 640–669 of eIF4GI. Binding was substantially improved when the eIF4GI fragment contained the eIF4E binding site and eIF4E was present in the binding assay. Lpro interacts with eIF4GI via residue C133 and residues 183–195 of the C‐terminal extension. This binding domain lies about 25 Å from the active site. Here, we examined the binding of Lpro to eIF4GI fragments generated by in vitro translation to narrow the binding site down to residues 645–657 of human eIF4GI. Comparison of these amino acids with those in human eIF4GII as well as with sequences of eIF4GI from other organisms allowed us to identify two conserved basic residues (K646 and R650). Mutation of these residues was severely detrimental to Lpro binding. Similarly, comparison of the sequence between residues 183 and 195 of Lpro with those of other FMDV serotypes and equine rhinitis A virus showed that acidic residues D184 and E186 were highly conserved. Substitution of these residues in Lpro significantly reduced eIF4GI binding and cleavage without affecting self‐processing. Thus, FMDV Lpro has evolved a domain that specifically recognizes a host cell protein.

Three-dimensional structure of foot-and-mouth disease virus and its biological functions

“…Meanwhile, residue Leu143 also determines the cleavage specificity of Lb pro [104]. Lb pro has a flexible C-terminal extension (CTE) of residues Asp184-Lys201, which are involved in self-processing and recognition of eIF4G [54,60]. The crystal structure of CTE in full-length Lb pro reveals that Lb pro appears to be a dimer because of the CTE folding back and interacting with the active site of a neighboring molecule, whereas that of sLb pro lacking six C-terminal amino acid residues is undefined because it cannot be traced in the electron density ( Fig.…”

The 3′ Untranslated Region of the Andes Hantavirus Small mRNA Functionally Replaces the Poly(A) Tail and Stimulates Cap-Dependent Translation Initiation from the Viral mRNA

“…Both possess similar biochemical activities and are functionally interchangeable (39). The FMDV L protease cleaves both eIF4GI and eIF4GII (13,15,46), inactivating the eIF4F complex in terms of its ability to recognize capped mRNAs, which results in a reduction of cap-dependent translation initiation (15,39). Interestingly, the carboxy-terminal fragment of eIF4G generated by FMDV L protease cleavage efficiently supports and indeed stimulates cap-independent translation initiation (9, 33).…”

Investigating the Substrate Specificity and Oligomerisation of the Leader Protease of Foot and Mouth Disease Virus using NMR

“…We have shown that Lb pro can contact the eIF4G homologues using a site away from the canonical substrate-binding cleft. 14,15 Nevertheless, the dimer still has to come apart so that the eIF4G substrate can enter the active site of Lb pro to allow proteolytic cleavage.…”

“…The CTE has been shown, together with Cys133, to have a role in recognising eIF4G, even though it is some 30 Å distant from the active site. 5,14,15 The CTE is clearly defined in the crystal structure of full-length Lb pro (PDB ID code 1QOL). The CTE of one molecule lies in the active site of an adjacent molecule and vice versa, 12 so that Lb pro appears as a dimer.…”