BACKBONE DYNAMICS OF THE A-DOMAIN OF HMG1 AS STUDIED BY N-15 NMR-SPECTROSCOPY

The HMG-box sequence motif (similar to 80 residues) occurs in a number of abundant eukaryotic chromosomal proteins such as HMG1, which binds DNA without sequence specificity, but with ''structure specificity'', as well as in several sequence-specific transcription factors. HMG1 h as two such boxes, A and B, which show similar to 30% sequence identit y, and an acidic C-terminal tail. The boxes are responsible for the ab ility of the protein to bend DNA and bind to bent or distorted DNA. Th e structure of the HMG box has been determined by NMR spectroscopy for the B-domain of HMG1 [Weir et al. (1993) EMBO J. 12, 1311-1319; Read et al. (1993) Nucleic Acids lies. 21, 3427-3436) and for Drosophila HM G-D (Jones et al. (1994) Structure 2, 609-627]. It has an unusual twis ted L-shape, suggesting that the protein might tumble anisotropically in solution. In this paper we report studies of the A-domain from HMG1 using N-15 NMR spectroscopy which show that the backbone dynamics of the protein can be described by two different rotational correlation t imes of 9.0 +/- 0.5 and 10.8 +/- 0.5 ns. We show that the relaxation d ata can be analyzed by assuming that the protein is a rigid, axially s ymmetric ellipsoid undergoing anisotropic rotational diffusion; the gl obal rotational diffusion constants, D-parallel to and D-perpendicular to, were estimated as 2.47 x 10(7) and 1.49 x 10(7) s(-1), respective ly. By estimating the angle between the amide bond vectors and the maj or axis of the rotational diffusion tensor from the family of structur es determined by NMR spectroscopy [see accompanying paper, Hardman et al. (1995) Biochemistry 34, 16596-16607], we were able to show that th e ellipsoid spectral density equation can reproduce the major features of the N-15 T-1 and T-2 profiles of the three helices in the HMG1 A-d omain. The backbone dynamics of the A-domain were then compared with t hose of the B-domain and the HMG box from HMG-D. This comparison stron gly supported the differences observed in the orientation of helix I i n the three structures, where the B-domain appears to be more similar to HMG-D than it is to the A-domain. These differences may turn out to be related to subtle differences in the DNA-binding properties of the A- and B-domains of HMG1.