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PMID: 8508775 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

DNA looping and the helical repeat in vitro and in vivo: effect of HU protein and enhancer location on Hin invertasome assembly.

The EMBO journal ·Vol. 12 ·No. 6 ·1993-06-00 ·Pages 2503-12

Haykinson MJ, Johnson RC

Abstract

Site-specific DNA inversion by the Hin recombinase requires the formation of a multicomponent nucleo-protein structure called an invertasome. In this structure, the two recombination sites bound by Hin are assembled together at the Fis-bound recombinational enhancer with the requisite looping of the intervening DNA segments. We have analyzed the role of the HU protein in invertasome assembly when the enhancer is located at variable positions close to one of the recombination sites. In the absence of HU in vitro and in hupA hupB mutant cells in vivo, invertasome assembly is very inefficient when there is < 104 bp of DNA between the enhancer and recombination site. Invertasome assembly in the presence of HU in vitro or in vivo displayed a periodicity beginning with 60 bp of intervening DNA that reflected its helical repeat. The average helical repeat for this DNA region was calculated by autocorrelation and Fourier transformation to be 11.2 bp per turn for supercoiled DNA both in the presence of HU in vitro and in hup+ cells in vivo. HU is the only protein in Escherichia coli that can promote invertasome formation with short DNA lengths between the enhancer and recombination sites. However, the presence of certain polyamines and a protein activity present in HeLa nuclear extracts can efficiently substitute for HU in invertasome assembly. These data support a model in which HU binds non-specifically to the DNA between the enhancer and recombination site to facilitate DNA looping.

MeSH Terms
Bacterial Proteins/chemistry,metabolism Base Sequence DNA Nucleotidyltransferases/metabolism DNA, Superhelical DNA-Binding Proteins/chemistry,metabolism Enhancer Elements, Genetic Escherichia coli/genetics,metabolism HeLa Cells Humans Molecular Sequence Data Nucleic Acid Conformation Polyamines/pharmacology Repetitive Sequences, Nucleic Acid
Chemicals
Bacterial Proteins DNA, Superhelical DNA-Binding Proteins Polyamines histone-like protein HU, bacteria DNA Nucleotidyltransferases Hin recombinase
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Haykinson M J
Department of Biological Chemistry, UCLA School of Medicine 90024-1737.
Johnson R C
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
Published
1993-06-00
Pages
2503-12
Language
English
Region
England
NLM ID
8208664
PMCID
PMC413488
Subset
IM
Grants
NIGMS NIH HHS · GM38509 · United States
External Links
PubMed source PMC Full Text View journal details
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