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Multiconformational states in phosphoglycerate dehydrogenase.
Jessica K Bell1, Gregory A Grant, Leonard J Banaszak
1Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Biochemistry
|March 24, 2004
Summary
Phosphoglycerate dehydrogenase (PGDH) regulation by serine involves cooperative binding. A W139G mutation disrupts this cooperativity, revealing structural flexibility and new insights into allosteric mechanisms in the serine biosynthetic pathway.
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- Phosphoglycerate dehydrogenase (PGDH) is key in serine biosynthesis.
- PGDH activity is allosterically regulated by serine in a cooperative manner.
- Previous structural studies revealed a tetrameric, toroidal PGDH structure.
Purpose of the Study:
- To investigate the role of residue W139 in PGDH's allosteric regulation.
- To characterize the structural and functional consequences of the W139G mutation.
- To understand the mechanism of serine-mediated V(max) regulation.
Main Methods:
- Site-directed mutagenesis to create W139G PGDH.
- Enzymatic assays to assess serine responsiveness and cooperativity.
- X-ray crystallography to determine the 2.09 Å structure of W139G-PGDH.
Main Results:
- The W139G mutant is enzymatically active but lacks cooperative serine inhibition.
- The crystal structure of W139G-PGDH shows significant quaternary and tertiary structural changes.
- Residues around W139 are conformationally flexible in the mutant, unlike the wild-type enzyme.
- A domain rotation of ~42 degrees was observed within subunits, leading to novel quaternary contacts.
Conclusions:
- Residue W139 is critical for relaying allosteric signals in PGDH.
- PGDH allosteric regulation involves conformational changes at the W139-containing interface, in addition to the ACT domain.
- The enzyme exhibits significant flexibility, allowing for novel structural conformations.