Purification and biosynthesis of a derepressible periplasmic arylsulfatase from Chlamydomonas reinhardtii.
The unicellular green alga Chlamydomonas reinhardtii responds to sulfate deprivation by producing an arylsulfatase (Lien, T., and O. Schreiner. 1975. Biochim. Biophys. Acta. 384:168-179; Schreiner, O., 1975. Biochim. Biophys. Acta. 384:180-193) and by developing the capacity to transport sulfate more rapidly (our unpublished data). The arylsulfatase activity, detectable 3 h after the transfer of the cells to low sulfate medium (less than or equal to 10 microM sulfate), is a periplasmic protein released into the culture medium by cw15, a cell wall-less mutant of C. reinhardtii. We have purified the derepressible arylsulfatase to homogeneity and have raised monospecific antibodies to it. The protein monomer (67.6 kD) associates into a dimer, and the enzyme activity shows an alkaline pH optimum and a Km of 0.3 mM for p-nitrophenylsulfate. Studies focused on arylsulfatase biosynthesis demonstrate that it is glycosylated and synthesized as a higher molecular mass precursor. The mature protein contains complex N-linked oligosaccharides and the primary translation product has an apparent molecular mass approximately 5 kD larger than the deglycosylated monomer. Since translatable RNA encoding the arylsulfatase can only be detected in cells after sulfate starvation, it is likely that accumulation of the enzyme is regulated at the level of transcription, although posttranscriptional processes may also be involved.
de Hostos
E L
EL
Department of Biological Sciences, Stanford University, California 94305.
Togasaki
R K
RK
Grossman
A
A
D001192:Arylsulfatases / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D002451:Cell Compartmentation; D011232:Chemical Precipitation; D002696:Chlamydomonas / Q000201:enzymology*; D002845:Chromatography; D004794:Enzyme Repression; D005786:Gene Expression Regulation; D006031:Glycosylation; D008970:Molecular Weight; D012333:RNA, Messenger / Q000235:genetics; D013429:Sulfatases / Q000302:isolation & purification*; D013455:Sulfur / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology
D012333:RNA, Messenger; D014415:Tunicamycin; D013455:Sulfur; D013429:Sulfatases; D001192:Arylsulfatases