MOLECULAR-INTERACTIONS BETWEEN THE HSV-1 CAPSID PROTEINS AS MEASURED BY THE YEAST 2-HYBRID SYSTEM

HSV-1 B capsids are composed of seven major proteins, designated VP5, VP19C, 21, 22a, VP23, VP24, and VP26. VP indicates that the capsid pro tein is also a component of the infectious virion. Capsid proteins 21, 22a, and VP24 are specified by a single open reading frame (UL26) tha t encodes 635 amino acids. An objective of the work in our laboratory is to identify and map interactions among and between capsid proteins. In the present studies we employed the yeast GAL4 two-hybrid system d eveloped by Fields and his colleagues (Nature 240, 245-246 (1989)) for this purpose. DNA corresponding to the capsid open reading frames was derived as a PCR product and fused to sequences of the GAL4 activatio n and DNA binding domains. Using this system each of the capsid protei ns has been tested for interactions with all of the other capsid prote ins. Three interactions have been identified: a relatively strong self -interaction between 22a molecules (residues 307-635 of UL26), bimolec ular interactions between 22a and VP5, and another between VP19C and V P23. The interactions were detected by the expression of beta-galactos idase enzyme activity, and yielded 289, 86, and 63 units of enzyme act ivity, respectively. For the 22a self-interaction, elimination of resi dues 611-635 resulted in an approximately twofold decrease in enzyme a ctivity. The C-terminal 25 amino acids of 22a were also essential for the bimolecular interaction between 22a and VF5. (C) 1996 Academic Pre ss, Inc.