Sequence organization and origin of HSV-1 strain Angelotti (ANG) class II defective DNA (HSV-1 ANG dDNA1) were examined in detail by establishing physical maps and by molecular cloning. dDNA1 consists of concatemers of tandem repeat units in which sequences from the U region spanning map coordinates 0.37 to 0.415 of standard HSV ANG DNA are covalently linked to TR/IR sequences. The size of the repeat unit was determined to be about 8.9 kilobase pairs (kb), comprising sequences of 7.3 kb from U and 1.6 kb from TR/IR regions. U sequences were delineated by restriction enzyme sites I N-P and RI F-M, and were colinear with the corresponding sequences of the standard (wild-type) virus genome. Expression of dDNA1 was studied in African green monkey kidney cells and in oocytes. A major polypeptide of approx. mol. wt. 135000 (135K) was overproduced, suggesting that this protein was encoded by dDNA1. By several parameters, e.g. size, immune cross-reactivity, and affinity for native and denatured DNA, the 135K polypeptide was identified as the major HSV DNA-binding protein. It was further shown that the repeat unit contains part of the DNA polymerase gene as demonstrated by its ability to rescue some mutations in this gene.

Keyword(s): defective DNA , HSV , mapping and polypeptides

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