J Gen Virol Faster Access
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via CrossRef
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Hang, X.
Right arrow Articles by Guarino, L. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Hang, X.
Right arrow Articles by Guarino, L. A.
Agricola
Right arrow Articles by Hang, X.
Right arrow Articles by Guarino, L. A.
Journal of General Virology (1999), 80, 2519-2526.
© 1999 Society for General Microbiology

Insect

Purification of Autographa californica nucleopolyhedrovirus DNA polymerase from infected insect cells

Xin Hang1 and Linda A. Guarino1

Departments of Biochemistry & Biophysics1 and Entomology2, Texas A&M University, College Station, TX 77843-2128, USA

Author for correspondence: Linda Guarino (at Department of Biochemistry & Biophysics). Fax +1 409 845 9274. e-mail lguarino{at}tamu.edu

Autographa californica nucleopolyhedrovirus (AcMNPV) DNA polymerase was purified from virus-infected cells using conventional chromatographic methods. The enzymatic activity of fractions eluting from single-stranded agarose gels was found to exactly coincide with a single polypeptide with an apparent molecular mass of approximately 110000 Da on denaturing polyacrylamide gels stained with Coomassie blue. This purification scheme resulted in a 228-fold purification of AcMNPV DNA polymerase with recovery of 3·5% of the initial activity. The specific activity of the most purified fraction of DNA polymerase was 5000 units/mg, which is sufficiently high to eliminate the possibility that contaminants significantly contribute to the polymerase activity. Preparations of purified DNA polymerase had 3'–5' exonuclease activity, but no 5'–3' exonuclease activity. The proofreading activity was apparently an intrinsic property of the enzyme as the ratio of nuclease activity to polymerase activity was constant throughout purification. Using a singly-primed M13 DNA template, RF-II DNA was detected within 3 min, indicating a polymerization rate of 40 nt/s. The effects of several DNA polymerase inhibitors on the enzymatic activity of purified DNA polymerase were also determined.




This article has been cited by other articles:


Home page
 J. Virol.Home page
E. A. Crouch and A. L. Passarelli
Genetic Requirements for Homologous Recombination in Autographa californica Nucleopolyhedrovirus
J. Virol., August 12, 2002; 76(18): 9323 - 9334.
[Abstract] [Full Text] [PDF]

Home page
 J. Gen. Virol.Home page
J. Huang and D. B. Levin
Expression, purification and characterization of the Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) DNA polymerase and interaction with the SpliNPV non-hr origin of DNA replication
J. Gen. Virol., July 1, 2001; 82(7): 1767 - 1776.
[Abstract] [Full Text] [PDF]

Home page
 J. Virol.Home page
V. V. McDougal and L. A. Guarino
The Autographa californica Nuclear Polyhedrosis Virus p143 Gene Encodes a DNA Helicase
J. Virol., June 1, 2000; 74(11): 5273 - 5279.
[Abstract] [Full Text]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
INT J SYST EVOL MICROBIOL MICROBIOLOGY J GEN VIROL
J MED MICROBIOL ALL SGM JOURNALS
Copyright © 1999 by the Society for General Microbiology.