J. Mol. Biol. (1990) 216. 141-160 o 2.9 A Resolution Structure of the N-terminal Domain of a Variant Surface Glycoprotein from Trypanosoma brucei Douglas Freymann~1 ", James Down ~, Mark Carrington 2, Isabel Roditi 3 Mervyn Turner 4 and Don Wiley ~ ~Department of Biochemistry and Molecular Biology and Howard Hughes Medical Institute Harvard University, 7 Divinity Avenue, Cambridge, MA 02138, U.S.A. "Department of Biochemistry, University of Cambridge Tennis Court Road, Cambridge CB2 1QW, U.K, aKernforschungzentrum Karlsruhe, Institut fiir Genetik und Toxikologie von Spaltstoffen Karlsruhe, Federal Republic of Germany ~Merck, Sharp and Dohme Research Laboratories P.O. Box 2000, Rahway, NJ 07065, U.S.A. (Received 11 May 1990; accepted 4 July 1990) The variant surface glyeoprotein (VSG) of Trypanosoma brucei forms a coat on the surface of the parasite; by the expression of a series of antigenically distinct VSGs in the surface coat the parasite escapes the host immune response. The 2.9 A resolution crystal structure of the N-terminal domain of one variant, MITat 1.2, has been determined. The structure was solved using data collected from two crystal forms. Initially a partial model was built into an electron density map based on multiple isomorphous replacement phases and improved by phase combination methods. Subsequently this model was used to obtain the molecular replacement solution tbr a second crystal form, providing starting phases which were refined using 2-fold non-crystallographic symmetry averaging. The current model includes 362 residues and has been refined using X-PLOR to an R value of 0.22 for data between 7 and 2-9 A. The molecule is a dimer, ~100A long, having an asymmetrical cross section with maximum dimensions of -~40 A × 60 A. Two long, ~70 A, a-helices from each monomer pack together to form, with several other helices, a core helix bundle that extends nearly the full length of the molecule. The "top" of the protein, which in the surface coat may be exposed to tile external environment, is formed from the ends of the two long helices, a short three-stranded/?-sheet, and a strand having irregular conformation that packs above these secondary structure elements. Two conserved disulfide bridges are in this part of the molecule. Several elements of the MITat 1.2 sequence, which contribute to the formation of the helix bundle structure, have been identified. These elements can be found in the sequences of several different VSGs, suggesting that to some extent the VSG structure is conserved in those variants. 1. Introduction The African trypanosome is a protozoan blood- stream parasite of mammMs which causes sleeping sickness in humans. Trypanosoma brucei is trans- mitted via an insect vector, the tsetse fly, being injected from the salivary gland when the fly takes a blood meal. Ill the bloodstream the parasite is t Present address: Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94143, U.S.A. 0022-2836/90/2OOl41-2o$03.00/0 covered by a surface coat, seen in electron micro- graphs as a 120 to 150 A (l A = 0"1 nm) thick elec- tron dense layer (Vickerman, 1969), composed of an estimated l0 T molecules of a single protein, the valiant surface glycoprotein (VSG:~: Cross, 1975). The trypanosome evades the immune response of :[:Abbreviations used: VSG, variant surface gtycoprotein; s.i.r., single isomorphous replacement; m.i.r., multiple isomorphous replacement; NCS, non-crystallographic symmetLy; r.m.s., root-mean-square. 141 © 1990 AcademicPrass Limited