Bluetongue virus core protein VP4 has nucleoside triphosphate phosphohydrolase activity.
The intact virion of bluetongue virus comprises ten segments of dsRNA enclosed in two concentric protein capsids. The core, which is transcriptionally active, includes three minor proteins (VP1, VP4 and VP6) which are considered to be the candidates for the core-associated enzymes that transcribe and modify full-length mRNA copies for each of the ten genome segments. Using purified recombinant VP4 protein and core-like particles containing VP4, in this report it is demonstrated that VP4 has nucleoside triphosphatase (NTPase) activity. VP4 is a nonspecific NTPase that hydrolyses four types of ribonucleoside triphosphate (NTP) to the corresponding nucleoside diphosphate. The substrate preference was GTP>ATP>UTP>CTP. NTP hydrolysis by VP4 was maximal when the Mg2+ or Ca2+ ion concentrations were 4 mM or 6 mM, respectively. The presence of single-stranded polynucleotides poly(A), poly(U) and poly(C) had little effect on the NTPase activity. Although the enzyme exhibited a broad temperature optimum around 40 degrees C, the pH optimum was sharp, between pH 7.5 and 8. The Km and Vmax of ATP hydrolysis were calculated to be 0.25+/-0.05 microM ATP and 55+/-4 pmol ATP hydrolysed min(-1) microg(-1), respectively. The Km was affected by the addition of poly(A) to only a small extent in contrast to the Vmax, which was increased by at least twofold.
Department of Biochemistry, University of Oxford, UK.
N
N
Ramadevi
P
P
Roy
D017766:Acid Anhydride Hydrolases / Q000032:analysis* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001820:Bluetongue virus / Q000737:chemistry*; D002118:Calcium / Q000494:pharmacology; D002213:Capsid / Q000032:analysis*; D036022:Capsid Proteins; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009696:Nucleic Acids / Q000494:pharmacology; D043583:Nucleoside-Triphosphatase; D018411:Spodoptera; D013379:Substrate Specificity; D013696:Temperature
D036022:Capsid Proteins; D009696:Nucleic Acids; C060339:VP4 protein, Rotavirus; D000255:Adenosine Triphosphate; D017766:Acid Anhydride Hydrolases; D043583:Nucleoside-Triphosphatase; D002118:Calcium