PROPERTIES OF A RECOMBINANT STRAIN OF HANSENULA-POLYMORPHA HAVING A TENFOLD INCREASED ACTIVITY OF DIHYDROXYACETONE KINASE

Biochemical and physiological properties were studied in the recombina nt of methylotrophic yeast strain Hansenula polymorpha pR22-2B grown i n a chemostat culture on medium with methanol. The strain contained 12 genome-integrated copies of a gene coding for homologous dihydroxyace tone kinase (DHAK) and had a tenfold increased activity of this enzyme . The enhanced activity of DHAK (the key enzyme of the methanol assimi lation pathway) in the recombinant strain leads to increased activitie s of methanol oxidase (the first enzyme of the methanol utilization pa thway) and formaldehyde dehydrogenase (the key enzyme of the methanol dissimilation pathway). This eventually results in increased efficienc y of formaldehyde utilization via its direct oxidation to CO2. These d ata imply that DHAK is indeed the rate-limiting step of the entire met hanol utilization pathway in H. polymorpha, and genetically engineered ''widening'' of this bottleneck results in increased efficiency of fo rmaldehyde oxidation to CO2, but does not enhance formaldehyde assimil ation into the biomass.