Separation and crystallization of T = 3 and T = 4 icosahedral complexes ofthe hepatitis B virus core protein

The icosahedral nucleocapsid of human hepatitis B virus is a homopolymer of the dimeric capsid protein also known as hepatitis B core antigen or HBcAg . Purified capsid protein obtained from an Escherichia coli expression syst em was reassembled into a mixture of T = 3 and T = 4 icosahedral particles consisting of 90 and 120 dimers, respectively. The two types of capsid were separated on a preparative scale by centrifugation through a sucrose gradi ent. In addition to this heterogeneity, the capsid protein has three cystei nes, one of which has a great propensity for forming disulfide bonds betwee n the two subunits, forming a dimer. To eliminate heterogeneity arising fro m oxidation, alanines were substituted for the cysteines. T = 3 and T = 4 c apsids crystallized under similar conditions. Crystals of T = 3 capsids dif fracted to similar to 8 Angstrom resolution; crystals of T = 4 capsids diff racted to 4 Angstrom resolution.