Synthesis of dolichol in a polyprenol reductase mutant is restored by elevation of cis-prenyl transferase activity.
CHB11-1-3 is a glycosylation mutant of Chinese hamster ovary (CHO) cells, isolated by screening mutagenized cells for those with decreased intracellular lysosomal enzyme activity [C. W. Hall et al. (1986) Mol. Cell. Biochem. 72, 35-45]. CHB11-1-3 synthesizes the lipid polyprenol, the metabolic precursor of dolichol, rather than dolichol, indicating a defect in polyprenol reductase. This defect was demonstrated previously in Lec9 CHO mutants, and cell fusion experiments confirmed that CHB11-1-3 is a member of this complementation group. A revertant of CHB11-1-3, CHBREV, isolated for its ability to grow at 39 degrees C, synthesizes dolichol at near-normal levels. CHBREV is probably a second-site revertant, because it synthesizes three to four times as much polyprenol as CHB11-1-3 and exhibits a similar elevation in the specific activity of cis-prenyl transferase. This higher activity appears to reflect an increase in enzyme molecules rather than the presence of an activator or absence of an inhibitor. These results suggest that CHB11-1-3 is a "K(m)" mutant, because synthesis of higher amounts of the substrate of polyprenol reductase obviates the defect.
Quellhorst
G J
GJ
Department of Biochemistry, The Johns Hopkins University, School of Hygiene and Public Health, Baltimore, Maryland 21205, USA.
Hall
C W
CW
Robbins
A R
AR
Krag
S S
SS
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004286:Dolichols / Q000096:biosynthesis*; D005816:Genetic Complementation Test; D016296:Mutagenesis; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D014166:Transferases / Q000378:metabolism*
D004286:Dolichols; D010088:Oxidoreductases; C106822:polyprenol reductase; D014166:Transferases; C106821:cis-prenyl transferase