The ROK1 gene was previously identified as a high-copy number plasmid
suppressor of the kem1 null mutation that affects microtubule and spin
dle pole body functions in Saccharomyces cerevisiae. The Rok1 protein
sequence contains highly conserved domains found in the DEAD protein f
amily of ATP-dependent RNA helicase. To investigate the crucial roles
of the RNA helicase domains found in the yeast ROK1 gene, we have intr
oduced amino acid substitution mutations in these regions. Two mutatio
ns were identified in the DEAD domain, which substitute the first Asp
with Glu. The D280E1 mutation, when subcloned onto the high-copy yeast
plasmid YEp24, no longer suppresses the slow growth phenotype and the
benomyl hypersensitivity of the kem1 mutation. Furthermore, the same
mutation does not complement the lethality caused by the rok1 null mut
ation. These results suggest that the DEAD helicase domain is crucial
to the ROK1 function.