Poliovirus RNA polymerase: in vitro enzymatic activities, fidelity of replication, and characterization of a temperature-sensitive RNA-negative mutant

Thesis/Dissertation · Tue Jan 01 00:00:00 EST 1985

OSTI ID:6941553

Stokes, M A.M.

The in vitro activities of the purified poliovirus RNA polymerase were investigated in this study. The polymerase was shown to be a strict RNA dependent RNA polymerase. It only copied RNA templates but used either a DNA or RNA primer to initiate RNA synthesis. Partially purified polymerase has some DNA polymerase activities. Additional purification of the enzyme and studies with a mutant poliovirus RNA polymerase indicated that the DNA polymerase activities were due to a cellular polymerase. The fidelity of RNA replication in vitro by the purified poliovirus RNA polymerase was studied by measuring the rate of misincorporation of noncomplementary ribonucleotide monophosphates on synthetic homopolymeric RNA templates. The results showed that the ratio of noncomplementary to complementary ribonucleotides incorporated was 1-5 x 10/sup -3/. The viral polymerase of a poliovirus temperature sensitive RNA-negative mutant, Ts 10, was isolated. This study confirmed that the mutant was viable 33/sup 0/, but was RNA negative at 39/sup 0/. Characterization of the Ts 10 polymerase showed it was significantly more sensitive to heat inactivation than was the old-type polymerase. Highly purified poliovirions were found to contain several noncapsid proteins. At least two of these proteins were labeled by (/sup 35/S)methionine infected cells and appeared to be virally encoded proteins. One of these proteins was immunoprecipitated by anti-3B/sup vpg/ antiserum. This protein had the approximate Mr = 50,000 and appeared to be one of the previously identified 3B/sup vpg/ precursor proteins.

• Research Organization: Florida Univ., Gainesville (USA)
• OSTI ID: 6941553
• Resource Relation: Other Information: Thesis (Ph. D.)
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
POLIO VIRUS
ENZYME ACTIVITY
GENETICS
RNA
BIOSYNTHESIS
RNA POLYMERASES
BIOCHEMISTRY
IN VITRO
METHIONINE
MUTANTS
PROTEINS
SULFUR 35
TEMPERATURE DEPENDENCE
AMINO ACIDS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGY
CARBOXYLIC ACIDS
CHEMISTRY
DAYS LIVING RADIOISOTOPES
DRUGS
ENZYMES
EVEN-ODD NUCLEI
ISOTOPES
LIGHT NUCLEI
LIPOTROPIC FACTORS
MICROORGANISMS
NUCLEI
NUCLEIC ACIDS
NUCLEOTIDYLTRANSFERASES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PARASITES
PHOSPHORUS-GROUP TRANSFERASES
POLYMERASES
RADIOISOTOPES
SULFUR ISOTOPES
SYNTHESIS
TRANSFERASES
VIRUSES
550701* - Microbiology- Tracer Techniques
550201 - Biochemistry- Tracer Techniques
550401 - Genetics- Tracer Techniques