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Nucleic Acids Research, 1986, Vol. 14, No. 2 945-963
© 1986

Articles

Identification, cloning and sequence determination of the genes specifying hexokinase A and B from yeast

Conrad Stachelek, Janet Stachelek, Judith Swan+, David Botstein+ and William Konigsberg

Department of Molecular Biophysics and Biochemistry, Yale University New Haven, CT 06510, USA +Department of Biology, Massachusetts Institute of Technology Cambridge, MA 02139, USA

Received November 14, 1985. Accepted December 13, 1985.

The hexokinase A (HKA) and hexokinase B (HKB) genes of Saccharomyces cerevisiae have been cloned from a library of yeast genomic DHA. Using an in vitro glucose phosphorylation assay, the HKB gene was located on a plasmid carrying a 13.6 kb fragment of yeast DHA. After subcloning the relevant restriction fragments, the nucleotide sequence of the HKB gene was determined. Using this information, we were able to locate the HKA gene on a plasmid carrying this gene, which we then seqoenced. Approximately 43% of the amino acid sequence of HKB was determined directly from 24 tryptic peptides. The results are in complete agreement with those derived from the DHA sequence and are consistent with the results of x-ray crystallography. Comparison of the amino acid sequences of HKA and HKB show that 378 out of 485 residues are identical. The 5' flanking region of the A gene contains nucleotide sequences expected for genes that are expressed at relatively high levels in yeast. The 24 base pair hyphenated palindrome at the 3' end of the HKB gene may be a site for termination of transcription of this gene.


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