JBC, Vol. 251, Issue 9, 2740-2743, May, 1976
Function and structure in phage Qbeta RNA replicase. Association of EF-Tu-Ts with the other enzyme subunits
T. Blumenthal, R. A. Young and S. Brown
Qbeta replicase is a complex of four nonidentical subunits readily
dissociable into two subcomplexes: 30 S ribosomal protein S1 and the
phage-coded polypeptide (Subunits I + II) and protein synthesis elongation
factors EF-Tu and EF-Ts (Subunits III + IV). The affinity of the two
subcomplexes for one another increases with increasing ionic strength. The
enzyme is capable of initiation of RNA synthesis with synthetic templates
only when in the low ionic strength conformation. Elongation of initiated
polynucleotide chains is not affectedby ionic strength. Addition of Qbeta
RNA to the enzyme also alters its quaternary structure: the EF-Tu-Ts cannot
be covalently attached to the other enzyme subunits with bifunctional
cross-linking reagents in the presence of RNA. This conformational change
is not influenced by ionic strength. The addition of Qbeta RNA to the
enzyme, does not result in the release of EF-Tu-Ts from the other enzyme
subunits: whereas free EF-Tu-Ts binds GDP independently of salt
concentration, this binding by Qbeta replicase is sensitive to high ionic
strength and remains so in the presence of Qbeta RNA. Furthermore, RNA does
not allow the release of EF-Ts from EF-Tu by GTP as measured by sensitivity
of EF-Ts activity to N-ethylmaleimide.