Electrostatic potential distribution of the gene V protein from Ff phage facilitates cooperative DNA binding: A model of the GVP‐ssDNA complex

Abstract

The crystal structure of the gene V protein (GVP) from the Ff filamentous phages (M13, f l , fd) has been solved for the wild-type and two mutant (Y41F and Y41H) proteins at high resolution. The Y41H mutant crystal structure revealed crystal packing interactions, which suggested a plausible scheme for constructing the polymeric protein shell of the GVP-single-stranded DNA (ssDNA) complex (Guan Y, et al., 1994, Biochemistry 33:7768-7778). The electrostatic potentials of the isolated and the cooperatively formed protein shell have been calculated using the program GRASP and they revealed a highly asymmetric pattern of the electrostatic charge distribution. The inner surface of the putative DNA-binding channel is positively charged, whereas the opposite outer surface is nearly neutral. The electrostatic calculation further demonstrated that the formation of the helical protein shell enhanced the asymmetry of the electrostatic distribution. A model of the GVP-ssDNA complex with the n = 4 DNA-binding mode could be built with only minor conformational perturbation to the GVP protein shell. The model is consistent with existing biochemical and biophysical data and provides clues to the properties of GVP, including the high cooperativity of the protein binding to ssDNA. The two antiparallel ssDNA strands form a helical ribbon with the sugar-phosphate backbones at the middle and the bases pointing away from each other. The bases are stacked and the Phe 73 residue is intercalated between two bases. The optimum binding to a tetranucleotide unit requires the participation of four GVP dimers, which may explain the cooperativity of the GVP binding to DNA.

Influence of the Compatible Solute Ectoine on the Local Water Structure: Implications for the Binding of the Protein G5P to DNA

“…The amount of bound G5P corrected for the bulk contribution is determined as 2702 RU (green line in Figure ), corresponding to an average number of 3.5 G5P molecules, which are directly interacting with each immobilized dT 25 oligonucleotide. This value can be compared with the expected one G5P molecule per four nucleotides, which has been reported in ref for mobile ssDNA in solution. The presence of a 1 M ectoine solution strongly reduces the amount of bound G5P to about one-third of the value obtained in the absence of ectoine.…”

“…In fact, structural studies of G5P 58,63 indicated the presence of highly charged regions which are of specific importance for the binding to DNA. Recent results also indicated a strong decrease of the dielectric constant of water in confined geometries, at interfaces and in electrolyte solutions ?…”

“…The importance of electrostatic interactions for proteins has also been recognized previously, , which might have some consequences for the binding of proteins to DNA. In fact, structural studies of G5P , indicated the presence of highly charged regions which are of specific importance for the binding to DNA. In combination, recent results also revealed a strong decrease of the dielectric constant of water in confined geometries, at interfaces, and in electrolyte solutions. ,, Thus, it can be assumed that these contributions in addition to the presence of ectoine might also have important consequences for the binding behavior.…”

Filamentous phages: masters of a microbial sharing economy

“…As the number of RFs templated for transcription increases, so too does the number of phage proteins present in the host cell. Late in the infection when the level of pV protein reaches a critical number, it forms dimers and binds at the bottom of the hairpin formed by the packaging signal in the IF DNA and begins to coat the entire length (except the packaging signal) of the DNA, with one antiparallel ssDNA strand bound by each side of the dimer forming a long helical DNA–protein complex containing approximately 1,600 copies of pV . pV also plays a role in timing the infection cycle in the host and coordinating the level of IF for packaging.…”

Optimizing protein V untranslated region sequence in M13 phage for increased production of single-stranded DNA for origami

“…Overall, one RF and (+) DNA are newly produced in each cycle from one RF DNA during stage 2. This stage continues until protein V (pV), a phage protein, accumulates to a threshold concentration ( 22 , 25 ). In stage 3, the RF DNA generated in stage 2 continues to replicate itself and generates one (+) strand DNA.…”