NDLI: Pre-steady-state DNA unwinding by bacteriophage T4 Dda helicase reveals a monomeric molecular motor

Content Provider	PubMed Central
Author	Bindu, Nanduri Byrd, Alicia K. Eoff, Robert L. Tackett, Alan J. Raney, Kevin D.
Copyright Year	2002
Abstract	Helicases are molecular motor enzymes that unwind and translocate nucleic acids. One of the central questions regarding helicase activity is whether the process of coupling ATP hydrolysis to DNA unwinding requires an oligomeric form of the enzyme. We have applied a pre-steady-state kinetics approach to address this question with the bacteriophage T4 Dda helicase. If a helicase can function as a monomer, then the burst amplitude in the pre-steady state might be similar to the concentration of enzyme, whereas if the helicase required oligomerization, then the amplitude would be significantly less than the enzyme concentration. DNA unwinding of an oligonucleotide substrate was conducted by using a Kintek rapid quench-flow instrument. The substrate consisted of 12 bp adjacent to 12 nucleotides of single-stranded DNA. Dda (4 nM) was incubated with substrate (16 nM) in buffer, and the unwinding reaction was initiated by the addition of ATP (5 mM) and Mg2+ (10 mM). The reaction was stopped by the addition of 400 mM EDTA. Product formation exhibited biphasic kinetics, and the data were fit to the equation for a single exponential followed by a steady state. The amplitude of the first phase was 3.5 ± 0.2 nM, consistent with a monomeric helicase. The burst amplitude of product formation was measured over a range of enzyme and substrate concentrations and remained consistent with a functional monomer. Thus, Dda can rapidly unwind oligonucleotide substrates as a monomer, indicating that the functional molecular motor component of a helicase can reside within a single polypeptide.
Related Links	http://dx.doi.org/10.1073/pnas.232401899
Ending Page	14727
Page Count	6
Starting Page	14722
File Format	PDF
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	23
Volume Number	99
Language	English
Publisher	National Academy of Sciences
Publisher Date	2002-11-12
Access Restriction	Open
Rights Holder	National Academy of Sciences
Subject Keyword	General Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

Sl.	Authority	Responsibilities	Communication Details
1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
5	Website/Portal (Helpdesk)	Queries regarding NDLI and its services	support@ndl.gov.in
6	Contents and Copyright Issues	Queries related to content curation and copyright issues	content@ndl.gov.in
7	National Digital Library of India Club (NDLI Club)	Queries related to NDLI Club formation, support, user awareness program, seminar/symposium, collaboration, social media, promotion, and outreach	clubsupport@ndl.gov.in
8	Digital Preservation Centre (DPC)	Assistance with digitizing and archiving copyright-free printed books	dpc@ndl.gov.in
9	IDR Setup or Support	Queries related to establishment and support of Institutional Digital Repository (IDR) and IDR workshops	idr@ndl.gov.in

Pre-steady-state DNA unwinding by bacteriophage T4 Dda helicase reveals a monomeric molecular motor

Measurement of steady-state kinetic parameters for DNA unwinding by the bacteriophage T4 Dda helicase: use of peptide nucleic acids to trap single-stranded DNA products of helicase reactions

Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism

Analysis of the bacteriophage T4 DNA polymerase holoenzyme and Dda helicase

Regulation of the Bacteriophage T4 Dda Helicase by Gp32 Single-stranded DNA-Binding Protein

Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism.

Stoichiometry and DNA Unwinding by the Bacteriophage T4 41:59 Helicase

Bacteriophage T4 Dda Helicase Translocates in a Unidirectional Fashion on Single-stranded DNA

Dda helicase tightly couples translocation on single-stranded DNA to unwinding of duplex DNA: Dda is an optimally active helicase

Pre-steady-state DNA unwinding by bacteriophage T4 Dda helicase reveals a monomeric molecular motor

Similar Documents

Pre-steady-state DNA unwinding by bacteriophage T4 Dda helicase reveals a monomeric molecular motor

Measurement of steady-state kinetic parameters for DNA unwinding by the bacteriophage T4 Dda helicase: use of peptide nucleic acids to trap single-stranded DNA products of helicase reactions

Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism

Analysis of the bacteriophage T4 DNA polymerase holoenzyme and Dda helicase

Regulation of the Bacteriophage T4 Dda Helicase by Gp32 Single-stranded DNA-Binding Protein

Real-time observation of bacteriophage T4 gp41 helicase reveals an unwinding mechanism.

Stoichiometry and DNA Unwinding by the Bacteriophage T4 41:59 Helicase

Bacteriophage T4 Dda Helicase Translocates in a Unidirectional Fashion on Single-stranded DNA

Dda helicase tightly couples translocation on single-stranded DNA to unwinding of duplex DNA: Dda is an optimally active helicase

Pre-steady-state DNA unwinding by bacteriophage T4 Dda helicase reveals a monomeric molecular motor