Foot-and-mouth disease virus leader proteinase: Structural insights into the mechanism of intermolecular cleavage

Abstract

Translation of foot-and-mouth disease virus RNA initiates at one of two start codons leading to the synthesis of two forms of leader proteinase Lpro (Labpro and Lbpro). These forms free themselves from the viral polyprotein by intra- and intermolecular self-processing and subsequently cleave the cellular eukaryotic initiation factor (eIF) 4G. During infection, Lbpro removes six residues from its own C-terminus, generating sLbpro. We present the structure of sLbpro bound to the inhibitor E64-R-P-NH2, illustrating how sLbpro can cleave between Lys/Gly and Gly/Arg pairs. In intermolecular cleavage on polyprotein substrates, Lbpro was unaffected by P1 or P1′ substitutions and processed a substrate containing nine eIF4GI cleavage site residues whereas sLbpro failed to cleave the eIF4GI containing substrate and cleaved appreciably more slowly on mutated substrates. Introduction of 70 eIF4GI residues bearing the Lbpro binding site restored cleavage. These data imply that Lbpro and sLbpro may have different functions in infected cells.

NMR analysis of the interaction of picornaviral proteinases Lb and 2A with their substrate eukaryotic initiation factor 4GII

“…For Lb pro , we used a naturally occurring shortened form (termed sLb pro ) lacking six amino acids at the C-terminus. 16,26 This deletion precludes protease dimerisation via interactions of the Cterminus of one molecule with the substrate binding site of a second and vice-versa, 27,28 thus simplifying complex formation and structural studies thereof. 29 For the 2A pro , we used wild-type HRV2 and CVB4 2A pro .…”

“…Structural information on the two proteinases as well as yeast and mammalian eIF4E is available. [14][15][16][17][18] For mammalian eIF4G, structural information [19][20][21] is limited to regions C-terminal to the cleavage sites of the picornaviral proteinases [ Fig. 1(A)]; nevertheless, the interaction of human eIF4E with human eIF4GI residues 557-646 has been characterised by isothermal titration calorimetry (ITC).…”

Structure and Function of Viral Deubiquitinating Enzymes

“…While a considerable amount of work has gone into the structural and functional characterization of L pro , a number of questions remain. While the autocatalytic activity and substrate specificity of L pro have been characterized, which have led to the development of peptidomimetic compounds as potential inhibitors of L pro [241][242][243], the molecular basis for Ub recognition by L pro remains to be elucidated. Mutagenesis has been able to discriminate between the DUB activity and polyprotein/ eIFG4 processing activities of L pro ; however, it is still unclear how these mutations exert their selective effect.…”

Picornavirus IRES elements: RNA structure and host protein interactions

“…The aphthovirus proteinase exists in two forms (designated Lab and Lb) derived from initiation of translation at either of two in-frame AUG codons (Cao et al, 1995). L pro is a papain-like protease (Guarne et al, 1998) that recognizes substrates rich in basic residues Steinberger et al, 2014).…”