Cloning and sequencing of the malate synthase gene from Hansenula polymorpha.
We have cloned the MAS gene, encoding the microbody matrix enzyme malate synthase (EC 4.1.3.2.) from the methylotrophic yeast Hansenula polymorpha. The gene was isolated by screening of a genomic library with a mixed-sequence probe, based on the partial amino acid sequence of the purified enzyme. The nucleotide sequence of a 2.4-kilobase stretch of DNA covering the MAS gene was determined. The gene contains an open reading frame of 555 amino acids, amounting to a calculated molecular mass of 63,254 for the encoded protein. Comparison of the amino acid sequence with the malate synthase sequences of Escherichia coli, Brassica napus L. and Cucumis sativus L. clearly establishes the homology of all four proteins. Compared to the soluble enzyme from E. coli, the malate synthases from H. polymorpha and both plant species, which are located in the microbodies, have a short carboxy-terminal extension. In the plant malate synthases, the extension is probably involved in routing to the microbodies, since it contains the potential peroxisomal targeting signal, Ser-Arg/Lys-Leu, at the carboxy terminus. The H. polymorpha enzyme terminates with similar amino acids, but their sequence, Ser-Leu-Lys, does not conform to any of the known peroxisomal targeting signals.
Bruinenberg
P G
PG
Laboratory of Biochemistry, Groningen University, The Netherlands.
Blaauw
M
M
Kazemier
B
B
Ab
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics*; D008292:Malate Synthase / Q000235:genetics*; D008830:Microbodies / Q000201:enzymology; D008969:Molecular Sequence Data; D007652:Oxo-Acid-Lyases / Q000235:genetics*; D010843:Pichia / Q000201:enzymology* / Q000235:genetics; D015183:Restriction Mapping; D004718:Saccharomycetales / Q000201:enzymology*
D004271:DNA, Fungal; D008292:Malate Synthase; D007652:Oxo-Acid-Lyases