Refined Structure of DNase 1 at 2 Å Resolution and Model for the Interaction with DNA
The structure of bovine pancreatic deoxyribonuclease I (DNase I) has been refined at 2 Å resolution using the restrained parameter LS refinement method of Hendrickson and Konnert. The conventional R-factor for the 16,104 reflections with I ??? 3 (I) from 6.0 to 2.0 Å resolution is 0.157. Bond lengths and angles of the refined structure are close to ideal values, the r.m.s. deviations being 0.023 Å and 1.4°, respectively. The r.m.s. deviation of short non bonded contacts from the sum of the van der Waals radii is 0.18 Å. The average atomic thermal vibration parameter, B, for the 2,031 atoms of the enzyme is 11.8 Å2. This low B-value is a consequence of the large hydrophobic core of DNase I formed by the two central, tightly packed ß-pleated sheets, which are surrounded by 8 helices and extended loop-regions. 377 water molecules have been located and refined, many of them are forming water-clusters. The protein sequence as determined chemically by the groups of Moore and Stein has to be corrected at 5 positions, the major correction being an insertion of 3 amino acid residues at R27. Two N-acetylglucosamine- and 5 mannose-residues have been located in the carbohydrate side-chain, which is branched at the mannose in position 3.
Based on the 3D-structure and the active site geometry determined from the binding of Ca2+-thymidine-3′, 5′-diphosphate in the crystal, we have derived a model for interactions of DNase I with ds-DNA. According to this model, the DNA binds at the slightly concave surface between the two ß-sheets with the exposed loop R70 — N71 — S72 — Y73 — K74 interacting in the minor groove of B-DNA. The hydroxyl group of Y73 is hydrogen-bonded to either 02 of a pyrimidine or N3 of a purine base. The positively charged residues R9, R70, R108, R38 and K74 are interacting with phosphates opposing each other in the minor groove. The cleavage patterns observed in oligonucleotides can be nicely explained in terms of this model.
KeywordsMinor Groove Groove Width Cleavage Pattern Refi Nement Shallow Groove
Unable to display preview. Download preview PDF.
- 4.A.V. Fratini, M.L. Kopka, H.R. Drew and R.E. Dickerson, J. Biol. Chem. 257:14686-14707.Google Scholar
- 6.W.A. Hendrickson and J.H. Konnert, In Computing in Crystallography (Diamond, R., Ramaseshan, S. and Venkatesan, K., eds.) pp. 13.01-13–32, Indian Academy of Sciences, Int. Union of Crystallography, Bangalore (1980).Google Scholar
- 10.M.S. Laskowski, in The Enzymes 3rd edition (Boyer, P.D., Ed.) Academic Press NY, Vol. IV, 289–311. (1971).Google Scholar
- 15.S. Moore, in The Enzymes 3rd edition (Boyer, P.D., Ed.) Academic Press NY, Vol. XIV, 281–296. (1981).Google Scholar