AN INTRAMOLECULAR DISULFIDE BRIDGE BETWEEN CYS-7 AND CYS61 DETERMINESTHE STRUCTURE OF THE SECRETORY CORE GENE-PRODUCT (E-ANTIGEN) OF HEPATITIS-B VIRUS

Hepatitis B vims, the prototypic member of the Hepadnaviridae, is a sm all enveloped DNA virus that replicates via reverse transcription. Eff icient usage of its compact 3.2-kb genome is exemplified by the pre-C/ C gene from which two proteins with largely overlapping primary sequen ces but distinctly different properties are synthesized: the self-asse mbling core protein p21c (hepatitis B core antigen [HbcAg]) and the se cretory, nonparticulate protein p17e (hepatitis B e antigen [HbeAg]). Mature p17e carries a 10-amino-acid N-terminal extension with a Cys re sidue (Cys-7). Using transient transfection of a human liver cell line with constructs expressing wild-type p17 or a series of Cys mutants o f p17, we show that Cys-7 forms an intramolecular S-S bond to Cys61, w hich in assembly-competent core proteins is available for intermolecul ar disulfide bonds between two neighboring subunits. Removal of the Cy s-7/Cys61 bond by mutating either residue has differential effects: in the absence of Cys-7, secretion is relatively efficient and independe nt of Cys61; however, the molecules are exported as homodimers exhibit ing both HBe and HBc antigenicity. In the absence of Cys61, the nonpai red Cys-7 interferes with secretion efficiency. The amino acid sequenc e flanking Cys-7 also contributes to the formation of the proper intra molecular S-S bond. These results suggest that the Cys-7/Cys61 bond im poses on p17e a conformation that is critical for its secretion and di stinct biophysical and antigenic properties. This mechanism adds selec tive disulfide formation to the repertoire of hepatitis B virus for ef ficient use of its tiny genome.