From owner-biophysics@net.bio.net Thu Jan 21 22:00:00 1999 From: "Iris Delmonte" Newsgroups: bionet.biophysics Subject: DNA Structure: Puzzle Number 20 Date: Fri, 22 Jan 1999 12:37:25 -0000 Lines: 55 X-Newsreader: Microsoft Outlook Express 4.72.3110.5 X-MimeOLE: Produced By Microsoft MimeOLE V4.72.3110.3 NNTP-Posting-Host: 195.7.225.221 Message-ID: <36a86b4e.0@news.netdirect.net.uk> X-Trace: 22 Jan 1999 12:13:02 GMT, 195.7.225.221 Path: biosci!agate!newsfeed.berkeley.edu!newsfeed.wirehub.nl!ayres.ftech.net!news.ftech.net!peer.news.nildram.co.uk!news-peer.netdirect.net.uk!news.netdirect.net.uk!195.7.225.221 Enhanced Rates of Cleavage of Duplex DNA Sakonju & Brown (47) reported on the contacts made by a 40,000 daltontranscription factor, tf IIIA, which protected its substrate from the actionof deoxyribonuclease 1 (RMM 31,000) between bases 44 to 90. Within that protected region, relative to bare duplex DNA, there is strong enhancement of cleavage at 60 & 61, and marked enhancement also at bases 74, 75 & 76, that is, 1.5 double helical turns away from 60 & 61. Since the two sites enjoy enhanced cleavage rates compared to bare DNA they must both be more accessible than is bare DNA, even in the middle of theprotected region, and even though tf III A must have some 2500 C,N,O atoms and the nuclease must have some 2000 C,N,O atoms. More specifically, site 60 & 61, if facing the nuclease, would leave site 74-76 on the opposite double helical face away from the nuclease, though both sites experience enhanced rates of cleavage compared to bare DNA and are only some 4.5 nm apart. Drew & Travers (48) record their frustration with this situation: "Many eukaryotic transcription factors, notably tf III A ... , protect up to 50 bp of DNA from enzymatic cleavage, yet somehow manage to induce regions of enhanced cleavage within their binding sites..." With the true side-by-side model (Puzzle 1, Ref.1) we can discern a possible explanation of these results. Tf III A may bind largely to the rear face where phosphates predominate, though, being a substantial protein, there may be some obstuction of the other face where the base pairs are stacked upon each other. This allows deoxyribonuclease 1 access to the stacked base pairs at both positions 60 - 61 and 1.5 turns away at 74 - 76, where both sites face way from tf III A, and the rates of cleavage are enhanced compared to bare DNA because the DNA bound onto tf III A is immobilised and entirely unfolded compared to its situation alone with the nuclease. No explanation of enhanced rates of cutting within zones of protection has ever been advanced based upon the double helical model of duplex DNA. 47 Contact Points between a Positive Transcription Factor and the Xenopus 5S RNA Gene; S. Sakonju & D.D. Brown; Cell Vol 31 (1982) 395 - 405 48 DNA Bending and its Relation to Nucleosome Positioning; H.R. Drew & A.A. Travers; J Mol Biol Vol 186 (1985) 773 - 790 --------------------------------------------------------------- Clive Delmonte .