Crystal structure of the coat protein from the GA bacteriophage: Model of the unassembled dimer

Abstract

There are four groups of RNA bacteriophages with distinct antigenic and physicochemical properties due to differences in surface residues of the viral coat proteins. Coat proteins also play a role as translational repressor during the viral life cycle, binding an RNA hairpin within the genome. In this study, the first crystal structure of the coat protein from a Group I1 phage GA is reported and compared to the Group I MS2 coat protein. The structure of the GA dimer was determined at 2.8 8, resolution (R-factor = 0.20). The overall folding pattern of the coat protein is similar to the Group I MS2 coat protein in the intact virus (Golmohammadi R, Valegird K, Fridborg K, Liljas L, 1993, J Mol Biol234:620-639) or as an unassembled dimer (Ni CZ, Syed R, Kodandapani R, Wickersham J, Peabody DS, Ely KR, 1995, Srructure 3:255-263). The structures differ in the FG loops and in the first turn of the a A helix. GA and MS2 coat proteins differ in sequence at 49 of 129 amino acid residues. Sequence differences that contribute to distinct immunological and physical properties of the proteins are found at the surface of the intact virus in the AB and FG loops. There are six differences in potential RNA contact residues within the RNA-binding site located in an antiparallel @sheet across the dimer interface. Three differences involve residues in the center of this concave site: Lys/Arg 83, Ser/Asn 87, and Asp/Glu 89. Residue 87 was shown by molecular genetics to define RNA-binding specificity by GA or MS2 coat protein (Lim F, Spingola M, Peabody DS, 1994, J BioZ Chem 269:9006-9010). This sequence difference reflects recognition of the nucleotide at position -5 in the unpaired loop of the translational operators bound by these coat proteins. In GA, the nucleotide at this position is a purine whereas in MS2, it is a pyrimidine.Keywords: bacteriophage coat protein; crystal structure; RNA hairpin; translational repressor; virus RNA bacteriophages are small viruses with a simple organization. These viruses have an icosahedral shell composed of 180 copies of coat protein and one copy of the maturation protein that encapsidate genomic RNA ,which is approximately 3,500 nucleotides in length. The single-stranded RNA genome also acts as mRNA, encoding four proteins: the viral coat protein, a maturation protein, a replicase subunit, and the lysis protein. These phages were first isolated from Escherichia coli, but later were also found in Caulobacter and Pseudomonas. To date, the coliphages have been the best studied and are classified into four groups based on serological and physicochemical properties. Groups I and I1 are quite similar, with MS2 and GA phages being the well-characterized members of these groups. Phages Q p and SP, members of Groups I11 and IV, respectively, are more divergent from Groups I and 11.

The crystal structure of bacteriophage GA and a comparison of bacteriophages belonging to the major groups of Escherichia coli leviviruses

“…A comparison of an AB H dimer from our capsid structure and the GA dimer crystal structure (Ni et al, 1996) shows that the root-mean-square (rms) deviation for C a carbon atoms is 1.2 A Ê . The largest deviation (6.8 A Ê ) is observed for AlaA96.…”

“…The main purpose of this work is to compare the structures of these phages and to try to conclude which features are the most important for the function and self assembly of the virus particle. The structure of a GA dimer has been recently established (Ni et al, 1996). The amino acid sequence of this protein differs at two positions from our sequence, and these differences seem to inhibit capsid formation.…”

In Vivo Packaging of mRNA in Yeast-Produced Bacteriophage GA Derived Virus-Like Particles

“…The first objective was to test GA capsid formation by expression of GA coat protein in yeast with three single amino acid mutations (amino acids 43, 55, 87) and with two double mutations (amino acids 43+87 or 55+87). Mutations for the introduction into GA coat protein sequence were chosen based on the analysis of already published data (Gott et al, 1991;Ni et al, 1996). It was also planned pIC921…”

The True Story and Advantages of RNA Phage Capsids as Nanotools

“…In addition to the structures of self-assembled RNA phage capsids, the crystal structures of unassembled mutant CP dimers of MS2 [100] and GA [62] have also been resolved. These structures showed only minor differences in comparison to their self-assembled counterparts.…”

“…The expression of RNA phage CP genes in E. coli led to the high-level production of correctly self-assembled icosahedral capsids that were morphologically and immunologically indistinguishable from virions in the case of MS2 [58,59] and fr [60] of group I, JP34, an intermediate between groups I and II [61], GA of group II [62], Qβ of group III [63], SP of group IV [64], the unclassified Levivirus members PP7 [65,66] and AP205 [67,68], and the unclassified Leviviridae phages PRR1 [69] and φCB5 [70]. It is noteworthy that not only E. coli cells but also the E. coli -based cell-free translation system can be used for efficient in vitro production of MS2 VLPs [71,72].…”