Macromolecular hydration changes associated with BamHI binding and catalysis.
In this report, the effects of osmotic pressure on BamHI cognate binding and catalysis were investigated and compared with a previous study on EcoRI (Robinson, C. R. and Sligar, S. G. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 2186-2191). Our observation of the dependence of binding and catalytic parameters on osmotic pressure has allowed for the comparison of hydration changes associated with site-specific DNA recognition for both endonucleases. Over a large range of osmotic pressures (pi), the dependence of BamHI on osmotic stress during cognate binding and catalysis was very different from that of the related endonuclease EcoRI. The binding of EcoRI to cognate DNA was dominated by a dehydration of the endonuclease-DNA complex, whereas binding by BamHI to its cognate sequence was accompanied by a solvent release corresponding to some 125 fewer waters. Catalytic analysis at elevated osmotic pressures indicated that both endonucleases had undergone a net hydration of the complex with BamHI displaying a much greater dependence on osmotic stress than EcoRI. Although the enzymes shared core structural motifs, comparisons of high resolution x-ray structures revealed many different secondary structural features of the complexed endonucleases. The large difference in hydration changes by both BamHI and EcoRI could be attributed to these dissimilar secondary structural features, as well as the functional differences of the two endonucleases during site-specific DNA recognition.
Lynch
T W
TW
Beckman Institute for Advanced Science and Technology and the Department of Biochemistry, University of Illinois, Urbana, Illinois 61801, USA.
Sligar
S G
SG
D001665:Binding Sites; D002384:Catalysis; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015245:Deoxyribonuclease BamHI / Q000378:metabolism*; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism; D009997:Osmotic Pressure; D011485:Protein Binding; D013816:Thermodynamics; D014867:Water / Q000378:metabolism*
D004268:DNA-Binding Proteins; D014867:Water; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI