Figure 2.

 Cloning and molecular analysis of the cut12⁺ gene. (a) Restriction mapping and complementation analysis of the cut12⁺ containing clones isolated by complementation of the cut12.1 mutant, and of subclones generated from the overlapping 2.8-kb region found in all cut12.1 complementing plasmids. (b) Nucleotide and predicted amino acid sequence of the cut12⁺ gene. The cut12⁺ gene contained 2 exons of 7 and 1637 nucleotides, which together encode a predicted 548 amino acid protein of 62 kD. The two exons are separated by an intron of 42 bp; the consensus intron splice and branch sequences are shown underlined and in italics. The two regions of the Cut12 protein predicted to form coiled coils are underlined. The potential p34^cdc2 and MAP kinase consensus sites are indicated in boldface type. (G+) The site of the stf1.1 mutation at glycine 71. The GenBank accession no. for the cut12⁺ sequence is Y16837. (c) Predicted coiled–coil regions in the Cut12 protein were determined by applying the algorithm of Lupas et al. (1991) with a 28 amino acid window. (d) Schematic representation of the main features of the Cut12 protein. (+) p34^cdc2 kinase consensus site; (star) MAP kinase consensus site; (hatched region) predicted region of coiled coil.