The energetics of HMG box interactions with DNA. Thermodynamic description of the box from mouse Sox-5.
The structural energetics of the HMG box from the DNA-binding protein mouse Sox-5 were examined calorimetrically. It was found that this box, notwithstanding its small size (molecular mass about 10 kDa), does not behave as a single cooperative unit and, on heating, the box reversibly unfolds in two separate stages. The first transition (tt approximately 34 degrees C) involves about 40% of the total enthalpy and the second (tt approximately 46 degrees C) the remainder. Both transitions proceed with significant heat capacity increment, showing that they are associated with the unfolding of two sub-domains having non-polar cores. According to heat capacity, ellipticity, fluorescence and NMR criteria, this HMG box is in a fully compact native state only below 5 degrees C. HMG boxes consist of two approximately orthogonal wings: the minor wing comprises helix 3 and its associated antiparallel N-terminal strand, whilst the major wing is composed of helices I and II. Analysis of the fluorescence and NMR spectra for this box obtained at different temperatures shows that the lower melting transition can be assigned to the minor wing and the upper transition to the major wing. Under physiological conditions (37 degrees C), the minor wing is considerably unfolded, whilst the major wing is essentially fully folded. DNA binding in vivo therefore involves refolding of the minor wing.
Department of Biology, Johns Hopkins University, Baltimore, MD 21218, USA.
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Crane-Robinson
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Read
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Cary
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Driscoll
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Dragan
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Privalov
D000818:Animals; D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006609:High Mobility Group Proteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008958:Models, Molecular; D009687:Nuclear Proteins / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D055755:SOXD Transcription Factors; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013816:Thermodynamics
D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; D009687:Nuclear Proteins; D055755:SOXD Transcription Factors; C089186:Sox5 protein, mouse; D004247:DNA