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Phosphatidylinositol synthase from yeast
1Department of Biochemical Engineering and Science, Faculty of Computer Science and Systems Engineering, Kyushu Institute of Technology, Iizuka, Fukuoka, Japan.
Biochimica Et Biophysica Acta
|November 25, 1997
Summary
Phosphatidylinositol (PI) synthase is crucial for yeast cell survival, synthesizing PI from CDP-diacylglycerol and myo-inositol. Its gene disruption is lethal, highlighting PI
Area of Science:
- Biochemistry
- Molecular Biology
- Yeast Genetics
Background:
- Phosphatidylinositol (PI) is vital for cellular functions, including lipid synthesis and protein anchoring.
- PI synthase synthesizes PI from CDP-diacylglycerol and myo-inositol, localized in yeast mitochondria and microsomes.
Purpose of the Study:
- To purify and characterize yeast PI synthase.
- To elucidate the enzyme's kinetic mechanism and regulation.
- To investigate the essentiality of PI synthase and PI in yeast.
Main Methods:
- Enzyme purification from yeast microsomes using Triton X-100.
- Enzyme activity assays with varying substrates and metal ions.
- Kinetic analysis to determine the reaction mechanism.
- Gene disruption and analysis of resulting phenotypes.
Main Results:
- PI synthase was purified and its activity found dependent on Mn2+/Mg2+ and Triton X-100.
- The enzyme follows a sequential Bi-Bi kinetic mechanism.
- PI synthase is a constitutive enzyme, insensitive to nutrient changes.
- A specific amino acid substitution (His114Gln) drastically increased Km for myo-inositol.
- Disruption of the PIS gene encoding PI synthase is lethal in yeast.
Conclusions:
- Yeast PI synthase is a constitutively expressed, essential enzyme with specific kinetic properties.
- The PIS gene and its encoded PI synthase are indispensable for yeast viability.
- Understanding PI synthesis provides insights into essential lipid metabolism pathways.