Effects of Poliovirus 3AB Protein on 3D Polymerase-catalyzed Reaction

Poliovirus RNA replication requires the activities of a viral RNA-dependent RNA polymerase, 3Dpol, in conjunction with several additional viral and likely cellular proteins. The importance of both the 3A and 3B coding regions has been documented previously by genetic tests, and their biochemical activities have been the subject of several recent investigations. In this study, we examined the previously reported stimulation of 3D-catalyzed RNA synthesis by 3AB. We show that 3AB does not stimulate RNA synthesis on templates that are stably base paired to a primer, indicating that 3AB does not stabilize or otherwise activate 3Dpol for chain elongation. Similarly, it does not alter the kinetic parameters or binding affinities of 3D for substrates. In the absence of a primer, or in the presence of a primer that does not form a stable hybrid with the template, 3AB increases the utilization of 3′-hydroxyl termini as sites for chain elongation by 3D, and thereby stimulates RNA synthesis. 3AB may interact with and stabilize these sites and/or may recruit 3Dpol to the site, resulting in stimulation of the initiation of elongation events. We propose that this activity is required for stabilizing weak interactions that occur during nucleotidyl-protein-primed initiation events in the viral RNA replication complex. Poliovirus RNA replication requires the activities of a viral RNA-dependent RNA polymerase, 3Dpol, in conjunction with several additional viral and likely cellular proteins. The importance of both the 3A and 3B coding regions has been documented previously by genetic tests, and their biochemical activities have been the subject of several recent investigations. In this study, we examined the previously reported stimulation of 3D-catalyzed RNA synthesis by 3AB. We show that 3AB does not stimulate RNA synthesis on templates that are stably base paired to a primer, indicating that 3AB does not stabilize or otherwise activate 3Dpol for chain elongation. Similarly, it does not alter the kinetic parameters or binding affinities of 3D for substrates. In the absence of a primer, or in the presence of a primer that does not form a stable hybrid with the template, 3AB increases the utilization of 3′-hydroxyl termini as sites for chain elongation by 3D, and thereby stimulates RNA synthesis. 3AB may interact with and stabilize these sites and/or may recruit 3Dpol to the site, resulting in stimulation of the initiation of elongation events. We propose that this activity is required for stabilizing weak interactions that occur during nucleotidyl-protein-primed initiation events in the viral RNA replication complex. Poliovirus-infected cells contain an RNA-dependent RNA polymerase activity that has been attributed to a virally encoded protein, now known as 3Dpol (1Lundquist R.E. Ehrenfeld E. Maizel J.V. Proc. Natl. Acad. Sci. U. S. A. 1974; 71: 4773-4777Crossref PubMed Scopus (34) Google Scholar, 2Flanegan J.B. Baltimore D. J. Virol. 1979; 29: 352-360Crossref PubMed Google Scholar, 3Kitamura N. Semler B.L. Rothberg P.G. Larsen G.R. Adler C.J. Dorner A.J. Emini E.A. Hanecak R. Lee J.J. Van der Werf S. Anderson C.W. Wimmer E. Nature. 1981; 291: 547-553Crossref PubMed Scopus (529) Google Scholar). A great deal of work over the past 25 years from many laboratories has been devoted to the purification and characterization of this enzyme, and several significant characteristics of 3Dpol and its biological activities have been determined. The poliovirus RNA polymerase is a 52-kDa protein that has been purified in large quantities to yield a protein with very few, if any, detectable contaminants (4Van Dyke T.A. Flanegan J.B. J. Virol. 1980; 35: 732-740Crossref PubMed Google Scholar, 5Plotch S.J. Palant O. Gluzman Y. J. Virol. 1989; 63: 216-225Crossref PubMed Google Scholar, 6Neufeld K.L. Richards O.C. Ehrenfeld E. J. Biol. Chem. 1991; 266: 24212-24219Abstract Full Text PDF PubMed Google Scholar). Detailed x-ray structural studies of a major portion of the protein have been completed (7Hansen J.L. Long A.M. Schultz S.C. Structure ( Lond. ). 1997; 5: 1109-1122Abstract Full Text Full Text PDF PubMed Scopus (380) Google Scholar) and establish the basic features common to all types of nucleic acid polymerases (DNA-directed as well as RNA-directed); these include “fingers,” “palm,” and “thumb” subdomains (8Kohlstaedt L.A. Wang J. Friedman J.M. Rice P.A. Steitz T.A. Science. 1992; 256: 1783-1790Crossref PubMed Scopus (1751) Google Scholar, 9Ollis D.L. Brick P. Hamlin R. Xuong N.G. Steitz T.A. Nature. 1985; 313: 762-766Crossref PubMed Scopus (736) Google Scholar). The enzyme is dependent on both a template and a primer for activity and is responsible for RNA chain elongation during poliovirus RNA replication in infected cells. Although the natural primer for RNA synthesis in poliovirus-infected cells remains to be definitively identified, synthetic primers are commonly used in the laboratory for assay and characterization of 3Dpol activity. It was proposed some years ago that structural elements in the RNA template may provide a mechanism for self-priming (10Tuschall D.M. Hiebert E. Flanegan J.B. J. Virol. 1982; 44: 209-216Crossref PubMed Google Scholar); however, a more current model involves protein priming (11Toyoda H. N. A. Wimmer E. J. Virol. PubMed Google Scholar). The enzyme has template by and it is known that in an infected 3Dpol viral protein, an in viral RNA as in the 3B coding may RNA synthesis Wimmer E. J. Virol. PubMed Google Scholar). It is on the of all RNA not completed Baltimore D. J. Virol. PubMed Google Scholar, N. Wimmer E. 5: PubMed Scopus Google Scholar, G.R. Dorner A.J. Wimmer E. 1980; PubMed Scopus Google indicating that the of or initiation of RNA synthesis. to the that may as primer for initiation of RNA synthesis. have been in infected cells by with Baltimore D. 1982; Full Text PDF PubMed Scopus Google Scholar, B.L. Anderson C.W. Hanecak R. Dorner Wimmer E. 1982; Full Text PDF PubMed Scopus Google Scholar, Semler B.L. Anderson C.W. Wimmer E. PubMed Scopus Google Scholar). are in the viral RNA replication from infected cells. a purified 3Dpol a and a RNA template, RNA was with and was with of this and the with RNA J. A. PubMed Scopus Google Scholar). that 3D and in this 3D to the of 3AB to of 3A a by synthesis of viral and/or a Baltimore D. J. Virol. PubMed Google Semler B.L. J. Virol. 1991; PubMed Google Scholar). It remains however, or some form as the of to RNA have that 3AB 3D activity in a J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). a template and primer was stimulation of in the presence of 3AB. the of both in the presence or absence of some large a RNA template with stimulation of 3D-catalyzed by 3AB was In a study, and Palant S.J. Palant O. J. Virol. PubMed Google Scholar) used poliovirus in a with a of reported that stimulation of 3D activity by 3AB with in the of chain to of 3AB on RNA synthesis by 3Dpol, reported 3AB activities (1Lundquist R.E. Ehrenfeld E. Maizel J.V. Proc. Natl. Acad. Sci. U. S. A. 1974; 71: 4773-4777Crossref PubMed Scopus (34) Google Scholar) 3AB to and 3D S.J. Palant O. J. Virol. PubMed Google A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). J.B. Baltimore D. J. Virol. 1979; 29: 352-360Crossref PubMed Google Scholar) 3AB in a with the the of poliovirus and the of Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). N. Semler B.L. Rothberg P.G. Larsen G.R. Adler C.J. Dorner A.J. Emini E.A. Hanecak R. Lee J.J. Van der Werf S. Anderson C.W. Wimmer E. Nature. 1981; 291: 547-553Crossref PubMed Scopus (529) Google Scholar) 3AB stimulates of to form 3D A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). (4Van Dyke T.A. Flanegan J.B. J. Virol. 1980; 35: 732-740Crossref PubMed Google Scholar) 3AB to RNA in a J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). a in 3AB RNA binding and RNA and synthesis in cells. S.J. Palant O. Gluzman Y. J. Virol. 1989; 63: 216-225Crossref PubMed Google Scholar) 3AB with as an protein Semler B.L. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google is not known or all of these activities significant to the RNA replication in an infected a biochemical mechanism for protein priming of RNA elongation has not been laboratories have reported the of of 3B both in and in that form base paired with the of poliovirus and and it is that these as primers for RNA synthesis. In this we show that 3AB does not stimulate 3D-catalyzed on templates that are stably base paired to a In the absence of primer in the presence of a primer that does not form a stable hybrid with the 3AB more utilization of 3′-hydroxyl termini that form 3AB may interact with and stabilize these sites and/or may recruit the 3D polymerase to the site, resulting in stimulation of the initiation of elongation events. We that this activity of 3AB initiation to occur on templates by weak interactions during nucleotidyl-protein-primed initiation of the and to the replication of poliovirus RNA is In the absence of a of the replication in several laboratories have studies of activities of and of known to be in RNA replication by genetic In this it is that be the replication in for the of 3AB in poliovirus RNA replication from of the by in the 3A or 3B coding from biochemical of 3AB with replication and from the presence of 3B on RNA several recent to the of 3AB on in the RNA replication it was that 3AB with RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) and with and 3D S.J. Palant O. J. Virol. PubMed Google Scholar, A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google J. Virol. 1997; 71: PubMed Google Scholar) and that it form a with and the termini of poliovirus RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Wimmer E. J. Virol. PubMed Google Scholar). In several have reported that 3AB stimulates 3D polymerase activity J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The biological of these to the viral RNA replication mechanism has to be this we have examined the stimulation of 3D-catalyzed RNA synthesis by 3AB in some We that 3AB does not stimulate 3D activity on a template that is stably base paired to a 3AB has on 3D or primer is or if the primer is not stably base paired to the template, 3AB stimulates 3D activity We that 3AB with 3′-hydroxyl that are base resulting in of a hybrid a for elongation. of 3D to the by in elongation activity. stimulation is 3D is interactions have a on 3D to the initiation 3D is in The that is responsible for the polymerase activity is with recent that 3AB stimulate polymerase activity by the RNA polymerase has on polymerase activity by on the template not with of to form stable with primer and in primer We have examined for as in the 3D in Although the used in very stable we to a stable hybrid the R. Baltimore D. 63: Full Text PDF PubMed Scopus Google Scholar) Similarly, stable not form from the of the poliovirus with from the and a of primers the with on is likely that used by J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google reported to significant stimulation of 3D activity by in a to the In a stimulation of the by 3AB J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). large stimulation may from hybrid on a template with large in 3D with 3AB have been previously J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). We have that these are from by and RNA polymerases from a of form hybrid for 3D of large be or be of and these be to with or RNA We have that with to by does not a of The large are in 3D 3AB are and is 3D does not of large hybrid that include the 3′-hydroxyl are not to 3D the used in the absence of by 3D on the was a to the of 3AB for a to the and is some and that both termini are not to the of in a template utilization is biochemical for 3AB in the of viral RNA replication in infected cells are its to as the for RNA initiation is with Baltimore D. 1982; Full Text PDF PubMed Scopus Google Scholar, B.L. Anderson C.W. Hanecak R. Dorner Wimmer E. 1982; Full Text PDF PubMed Scopus Google Scholar, Semler B.L. Anderson C.W. Wimmer E. PubMed Scopus Google Scholar) may the in the replication complex. to or not the interactions are may of 3AB on the The activity in this may in of the weak hybrid the template and a form of 3AB or for and stimulation of to 3D or binding and of 3D the polymerase and thereby chain initiation to be of 3AB chain elongation is studies be required to the importance of 3AB activities in the RNA replication Poliovirus-infected cells contain an RNA-dependent RNA polymerase activity that has been attributed to a virally encoded protein, now known as 3Dpol (1Lundquist R.E. Ehrenfeld E. Maizel J.V. Proc. Natl. Acad. Sci. U. S. A. 1974; 71: 4773-4777Crossref PubMed Scopus (34) Google Scholar, 2Flanegan J.B. Baltimore D. J. Virol. 1979; 29: 352-360Crossref PubMed Google Scholar, 3Kitamura N. Semler B.L. Rothberg P.G. Larsen G.R. Adler C.J. Dorner A.J. Emini E.A. Hanecak R. Lee J.J. Van der Werf S. Anderson C.W. Wimmer E. Nature. 1981; 291: 547-553Crossref PubMed Scopus (529) Google Scholar). A great deal of work over the past 25 years from many laboratories has been devoted to the purification and characterization of this enzyme, and several significant characteristics of 3Dpol and its biological activities have been determined. The poliovirus RNA polymerase is a 52-kDa protein that has been purified in large quantities to yield a protein with very few, if any, detectable contaminants (4Van Dyke T.A. Flanegan J.B. J. Virol. 1980; 35: 732-740Crossref PubMed Google Scholar, 5Plotch S.J. Palant O. Gluzman Y. J. Virol. 1989; 63: 216-225Crossref PubMed Google Scholar, 6Neufeld K.L. Richards O.C. Ehrenfeld E. J. Biol. Chem. 1991; 266: 24212-24219Abstract Full Text PDF PubMed Google Scholar). Detailed x-ray structural studies of a major portion of the protein have been completed (7Hansen J.L. Long A.M. Schultz S.C. Structure ( Lond. ). 1997; 5: 1109-1122Abstract Full Text Full Text PDF PubMed Scopus (380) Google Scholar) and establish the basic features common to all types of nucleic acid polymerases (DNA-directed as well as RNA-directed); these include “fingers,” “palm,” and “thumb” subdomains (8Kohlstaedt L.A. Wang J. Friedman J.M. Rice P.A. Steitz T.A. Science. 1992; 256: 1783-1790Crossref PubMed Scopus (1751) Google Scholar, 9Ollis D.L. Brick P. Hamlin R. Xuong N.G. Steitz T.A. Nature. 1985; 313: 762-766Crossref PubMed Scopus (736) Google Scholar). The enzyme is dependent on both a template and a primer for activity and is responsible for RNA chain elongation during poliovirus RNA replication in infected cells. Although the natural primer for RNA synthesis in poliovirus-infected cells remains to be definitively identified, synthetic primers are commonly used in the laboratory for assay and characterization of 3Dpol activity. It was proposed some years ago that structural elements in the RNA template may provide a mechanism for self-priming (10Tuschall D.M. Hiebert E. Flanegan J.B. J. Virol. 1982; 44: 209-216Crossref PubMed Google Scholar); however, a more current model involves protein priming (11Toyoda H. N. A. Wimmer E. J. Virol. PubMed Google Scholar). The enzyme has template by and it is known that in an infected 3Dpol viral The protein, an in viral RNA as in the 3B coding may RNA synthesis Wimmer E. J. Virol. PubMed Google Scholar). It is on the of all RNA not completed Baltimore D. J. Virol. PubMed Google Scholar, N. Wimmer E. 5: PubMed Scopus Google Scholar, G.R. Dorner A.J. Wimmer E. 1980; PubMed Scopus Google indicating that the of or initiation of RNA synthesis. to the that may as primer for initiation of RNA synthesis. have been in infected cells by with Baltimore D. 1982; Full Text PDF PubMed Scopus Google Scholar, B.L. Anderson C.W. Hanecak R. Dorner Wimmer E. 1982; Full Text PDF PubMed Scopus Google Scholar, Semler B.L. Anderson C.W. Wimmer E. PubMed Scopus Google Scholar). are in the viral RNA replication from infected cells. a purified 3Dpol a and a RNA template, RNA was with and was with of this and the with RNA J. A. PubMed Scopus Google Scholar). that 3D and in this 3D to the of 3AB to of 3A a by synthesis of viral and/or a Baltimore D. J. Virol. PubMed Google Semler B.L. J. Virol. 1991; PubMed Google Scholar). It remains however, or some form as the of to RNA have that 3AB 3D activity in a J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). a template and primer was stimulation of in the presence of 3AB. the of both in the presence or absence of some large a RNA template with stimulation of 3D-catalyzed by 3AB was In a study, and Palant S.J. Palant O. J. Virol. PubMed Google Scholar) used poliovirus in a with a of reported that stimulation of 3D activity by 3AB with in the of chain elongation. In to of 3AB on RNA synthesis by 3Dpol, reported 3AB activities (1Lundquist R.E. Ehrenfeld E. Maizel J.V. Proc. Natl. Acad. Sci. U. S. A. 1974; 71: 4773-4777Crossref PubMed Scopus (34) Google Scholar) 3AB to and 3D S.J. Palant O. J. Virol. PubMed Google A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). J.B. Baltimore D. J. Virol. 1979; 29: 352-360Crossref PubMed Google Scholar) 3AB in a with the the of poliovirus and the of Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). N. Semler B.L. Rothberg P.G. Larsen G.R. Adler C.J. Dorner A.J. Emini E.A. Hanecak R. Lee J.J. Van der Werf S. Anderson C.W. Wimmer E. Nature. 1981; 291: 547-553Crossref PubMed Scopus (529) Google Scholar) 3AB stimulates of to form 3D A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). (4Van Dyke T.A. Flanegan J.B. J. Virol. 1980; 35: 732-740Crossref PubMed Google Scholar) 3AB to RNA in a J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). a in 3AB RNA binding and RNA and synthesis in cells. S.J. Palant O. Gluzman Y. J. Virol. 1989; 63: 216-225Crossref PubMed Google Scholar) 3AB with as an protein Semler B.L. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). It is not known or all of these activities significant to the RNA replication in an infected a biochemical mechanism for protein priming of RNA elongation has not been laboratories have reported the of of 3B both in and in that form base paired with the of poliovirus and and it is that these as primers for RNA synthesis. In this we show that 3AB does not stimulate 3D-catalyzed on templates that are stably base paired to a In the absence of primer in the presence of a primer that does not form a stable hybrid with the 3AB more utilization of 3′-hydroxyl termini that form 3AB may interact with and stabilize these sites and/or may recruit the 3D polymerase to the site, resulting in stimulation of the initiation of elongation events. We that this activity of 3AB initiation to occur on templates by weak interactions during nucleotidyl-protein-primed initiation events. of the and to the replication of poliovirus RNA is In the absence of a of the replication in several laboratories have studies of activities of and of known to be in RNA replication by genetic In this it is that be the replication in for the of 3AB in poliovirus RNA replication from of the by in the 3A or 3B coding from biochemical of 3AB with replication and from the presence of 3B on RNA several recent to the of 3AB on in the RNA replication it was that 3AB with RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) and with and 3D S.J. Palant O. J. Virol. PubMed Google Scholar, A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google J. Virol. 1997; 71: PubMed Google Scholar) and that it form a with and the termini of poliovirus RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Wimmer E. J. Virol. PubMed Google Scholar). In several have reported that 3AB stimulates 3D polymerase activity J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The biological of these to the viral RNA replication mechanism has to be this we have examined the stimulation of 3D-catalyzed RNA synthesis by 3AB in some We that 3AB does not stimulate 3D activity on a template that is stably base paired to a 3AB has on 3D or primer is or if the primer is not stably base paired to the template, 3AB stimulates 3D activity We that 3AB with 3′-hydroxyl that are base resulting in of a hybrid a for elongation. of 3D to the by in elongation activity. stimulation is 3D is interactions have a on 3D to the initiation 3D is in The that is responsible for the polymerase activity is with recent that 3AB stimulate polymerase activity by the RNA polymerase has on polymerase activity by on the template not with of to form stable with primer and in primer We have examined for as in the 3D in Although the used in very stable we to a stable hybrid the R. Baltimore D. 63: Full Text PDF PubMed Scopus Google Scholar) Similarly, stable not form from the of the poliovirus with from the and a of primers the with on is likely that used by J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google reported to significant stimulation of 3D activity by in a to the In a stimulation of the by 3AB J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). large stimulation may from hybrid on a template with large in 3D with 3AB have been previously J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). We have that these are from by and RNA polymerases from a of form hybrid for 3D of large be or be of and these be to with or RNA We have that with to by does not a of The large are in 3D 3AB are and is 3D does not of large hybrid that include the 3′-hydroxyl are not to 3D the used in the absence of by 3D on the was a to the of 3AB for a to the and is some and that both termini are not to the of in a template utilization is biochemical for 3AB in the of viral RNA replication in infected cells are its to as the for RNA initiation is with Baltimore D. 1982; Full Text PDF PubMed Scopus Google Scholar, B.L. Anderson C.W. Hanecak R. Dorner Wimmer E. 1982; Full Text PDF PubMed Scopus Google Scholar, Semler B.L. Anderson C.W. Wimmer E. PubMed Scopus Google Scholar) may the in the replication complex. to or not the interactions are may of 3AB on the The activity in this may in of the weak hybrid the template and a form of 3AB or for and stimulation of to 3D or binding and of 3D the polymerase and thereby chain initiation to be of 3AB chain elongation is studies be required to the importance of 3AB activities in the RNA replication of the and to the replication of poliovirus RNA is In the absence of a of the replication in several laboratories have studies of activities of and of known to be in RNA replication by genetic In this it is that be the replication in for the of 3AB in poliovirus RNA replication from of the by in the 3A or 3B coding from biochemical of 3AB with replication and from the presence of 3B on RNA several recent to the of 3AB on in the RNA replication it was that 3AB with RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) and with and 3D S.J. Palant O. J. Virol. PubMed Google Scholar, A. J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google J. Virol. 1997; 71: PubMed Google Scholar) and that it form a with and the termini of poliovirus RNA Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, Wimmer E. J. Virol. PubMed Google Scholar). In several have reported that 3AB stimulates 3D polymerase activity J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google Scholar, J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The biological of these to the viral RNA replication mechanism has to be determined. In this we have examined the stimulation of 3D-catalyzed RNA synthesis by 3AB in some We that 3AB does not stimulate 3D activity on a template that is stably base paired to a 3AB has on 3D or primer is or if the primer is not stably base paired to the template, 3AB stimulates 3D activity We that 3AB with 3′-hydroxyl that are base resulting in of a hybrid a for elongation. of 3D to the by in elongation activity. stimulation is 3D is interactions have a on 3D to the initiation 3D is in The that is responsible for the polymerase activity is with recent that 3AB stimulate polymerase activity by the RNA polymerase has on polymerase activity by on the template not with of to form stable with primer and in primer We have examined for as in the 3D in Although the used in very stable we to a stable hybrid the R. Baltimore D. 63: Full Text PDF PubMed Scopus Google Scholar) Similarly, stable not form from the of the poliovirus with from the and a of primers the with on It is likely that used by J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, S.J. Palant O. J. Virol. PubMed Google reported to significant stimulation of 3D activity by in a to the In a stimulation of the by 3AB J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). large stimulation may from hybrid on a template with The large in 3D with 3AB have been previously J. Wimmer E. J. Biol. Chem. Full Text PDF PubMed Google Scholar). We have that these are from by and RNA polymerases from a of form hybrid for 3D of large be or be of and these be to with or RNA We have that with to by does not a of The large are in 3D 3AB are and is 3D does not of large hybrid that include the 3′-hydroxyl are not to 3D the used in the absence of 3AB. The by 3D on the was a to the of 3AB for a to the and is some and that both termini are not to the of in a template utilization is The biochemical for 3AB in the of viral RNA replication in infected cells are its to as the for RNA initiation is with Baltimore D. 1982; Full Text PDF PubMed Scopus Google Scholar, B.L. Anderson C.W. Hanecak R. Dorner Wimmer E. 1982; Full Text PDF PubMed Scopus Google Scholar, Semler B.L. Anderson C.W. Wimmer E. PubMed Scopus Google Scholar) may the in the replication complex. to or not the interactions are may of 3AB on the The activity in this may in of the weak hybrid the template and a form of 3AB or for and stimulation of to 3D or binding and of 3D the polymerase and thereby chain initiation to be of 3AB chain elongation is studies be required to the importance of 3AB activities in the RNA replication We for in of We and for on the