NEGATIVE EFFECT OF SEQUENTIAL SERINE CODONS ON EXPRESSION OF FOREIGN GENES IN ESCHERICHIA-COLI

Herpes simplex virus encodes a 1298-residue protein designated ICP4 th at regulates transcription of viral genes. Structural and functional a nalyses of ICP4 have been facilitated by production of portions of ICP 4 in Escherichia coli. We previously observed that expression of most truncated forms of ICP4 in E. coli was relatively efficient, with the exception of portions of the ICP4 gene approximately between codons 16 0 and 220. We have now localized the portion of ICP4 that inhibits exp ression to a serine-rich region from position 176 to 199. Our experime ntal results suggest that codons within the serine-rich domain do not induce termination of transcription, do not alter the intrinsic stabil ity of mRNA, and do not create a proteolytically sensitive site in thi s portion of ICP4. Silent mutations that alter codon usage of many of the 19 serine codons in this region had no effect on expression. Howev er, we observed that the level of protein expression was inversely pro portional to the number of serine codons in this region. The results a re consistent with a model in which the serine-rich domain induces pre mature termination of translation. This effect is not due to any speci fic secondary structure in the mRNA or lack of sufficient seryl-tRNA s ynthetase. It remains to be determined whether premature termination c an result from insufficient seryl-charged tRNAs. Our results suggest t hat foreign genes with more than 20 consecutive serine codons may be p oorly expressed in E. coli. (C) 1996 Academic Press, Inc.