A directed approach to improving the solubility of Moloney murine leukemiavirus reverse transcriptase

One of the difficulties that can impede structural work on a molecule of in terest is limited solubility. Although functionally similar to the human im munodeficiency virus type-1 reverse transcriptase (HIV-1 RT), the Moloney m urine leukemia virus reverse transcriptase (MMLV RT) differs both in archit ecture and solubility properties. Reverse transcriptase is an essential ret roviral enzyme that replicates the single-stranded RNA genome of the retrov irus producing a double-stranded DNA copy, which is subsequently integrated into the host's genome. We have introduced a single amino acid substitutio n in the connection domain of an N-terminally truncated MMLV RT (L435K) tha t significantly improves the solubility of the enzyme eliminating the need for nonionic detergents in buffering storage solutions. The substituted enz yme retains near wild-type polymerase activity. An important consequence of the improved solubility of the L435K MMLV RT has been the ability to obtai n diffraction quality crystals.