Steven S. Broyles

Bio: Steven S. Broyles is an academic researcher from Anschutz Medical Campus. The author has contributed to research in topics: DNA supercoil & Cruciform. The author has an hindex of 2, co-authored 3 publications receiving 404 citations.

Perfect palindromic lac operator DNA sequence exists as a stable cruciform structure in supercoiled DNA in vitro but not in vivo

Abstract: A perfect palindromic 66-base pair (bp) DNA sequence derived from the lac operator and cloned into plasmid pMB9 [Betz, J. L. & Sadler, J. R. (1981) Gene 13, 1-12] can exist in a 66-bp linear form or as two 33-bp cruciform arms. The fraction of the sequence in the cruciform depends on the superhelical density of the plasmid DNA. Relaxed DNA contains no cruciforms. The palindrome in the cruciform structure is cut by EcoRI endonuclease at the base of the cruciform arms, releasing 33-bp fragments; when in the linear form only 66-bp fragments are produced. The cruciform structure is fixed by trimethylpsoralen crosslinks in the cruciform arms. This together with the EcoRI cutting provides an assay for the cruciform structures in the DNA of living cells. Using this assay we show that the cruciform structure rarely if ever exists in vivo, but after DNA isolation greater than 90% of the sequence is in cruciforms. Results suggest that the plasmid DNA as organized in vivo either lacks sufficient torsional tension to form this cruciform or the palindrome is restrained in the linear form by other bound molecules.

Cruciform Transitions Assayed Using a Psoralen Crosslinking Method: Applications to measurements of DNA torsional tension

Abstract: It is now established that cruciform structures can form in supercoiled DNA at sites of inverted repeat sequences. Several different methods have been developed for detecting cruciforms, including: scission of the DNA with nucleases specific for the unpaired bases at the termini of cruciform arms (Lilley, 1980, 1981; Panayotatos et al; Singleton et al,); scission with restriction enzymes (Geliert et al; Courey et al); electrophoretic resolution of DNA containing or not containing a cruciform (Mizuuchi et al; Geliert et al); electron microscopic observation (Mizuuchi et al).

DNA Structure and Function

Abstract: Publisher Summary This chapter discusses the structure and function of DNA. DNA occupies a critical role in cells, because it is the source of all intrinsic genetic information. Chemically, DNA is a very stable molecule, a characteristic important for a macromolecule that may have to persist in an intact form for a long period of time before its information is accessed by the cell. Although DNA plays a critical role as an informational storage molecule, it is by no means as unexciting as a computer tape or disk drive. The structure of the DNA described by Watson and Crick in 1953 is a right handed helix of two individual antiparallel DNA strands. Hydrogen bonds provide specificity that allows pairing between the complementary bases (A.T and G.C) in opposite strands. Base stacking occurs near the center of the DNA helix and provides a great deal of stability to the helix (in addition to hydrogen bonding). The sugar and phosphate groups form a “backbone” on the outside of the helix. There are about 10 base pairs (bp) per turn of the double helix.

Bacterial nucleoid-associated proteins, nucleoid structure and gene expression

Abstract: Nucleoid-associated proteins (NAPs) bind to the bacterial chromosome and alter its dynamics, maintaining nucleoid structure. In this Review, Dillon and Dorman examine the range of proteins in the ever-growing NAP family and their contributions to the regulation of nucleoid structure and gene expression. Emerging models of the bacterial nucleoid show that nucleoid-associated proteins (NAPs) and transcription contribute in combination to the dynamic nature of nucleoid structure. NAPs and other DNA-binding proteins that display gene-silencing and anti-silencing activities are emerging as key antagonistic regulators of nucleoid structure. Furthermore, it is becoming clear that the boundary between NAPs and conventional transcriptional regulators is quite blurred and that NAPs facilitate the evolution of novel gene regulatory circuits. Here, NAP biology is considered from the standpoints of both gene regulation and nucleoid structure.