3HL4: Crystal Structure Of A Mammalian Ctp:phosphocholine Cytidylyltransferase With Cdp-Choline

Citation:
Abstract
CTP:phosphocholine cytidylyltransferase (CCT) is the key regulatory enzyme in the synthesis of phosphatidylcholine, the most abundant phospholipid in eukaryotic cell membranes. The CCT-catalyzed transfer of a cytidylyl group from CTP to phosphocholine to form CDP-choline is regulated by a membrane lipid-dependent mechanism imparted by its C-terminal membrane binding domain. We present the first analysis of a crystal structure of a eukaryotic CCT. A deletion construct of rat CCTalpha spanning residues 1-236 (CCT236) lacks the regulatory domain and as a result displays constitutive activity. The 2.2-A structure reveals a CCT236 homodimer in complex with the reaction product, CDP-choline. Each chain is composed of a complete catalytic domain with an intimately associated N-terminal extension, which together with the catalytic domain contributes to the dimer interface. Although the CCT236 structure reveals elements involved in binding cytidine that are conserved with other members of the cytidylyltransferase superfamily, it also features nonconserved active site residues, His-168 and Tyr-173, that make key interactions with the beta-phosphate of CDP-choline. Mutagenesis and kinetic analyses confirmed their role in phosphocholine binding and catalysis. These results demonstrate structural and mechanistic differences in a broadly conserved protein fold across the cytidylyltransferase family. Comparison of the CCT236 structure with those of other nucleotidyltransferases provides evidence for substrate-induced active site loop movements and a disorder-to-order transition of a loop element in the catalytic mechanism.
PDB ID: 3HL4Download
MMDB ID: 76874
PDB Deposition Date: 2009/5/26
Updated in MMDB: 2009/12
Experimental Method:
x-ray diffraction
Resolution: 2.2  Å
Source Organism:
Similar Structures:
Biological Unit for 3HL4: dimeric; determined by author and by software (PISA)
Molecular Components in 3HL4
Label Count Molecule
Proteins (2 molecules)
2
Choline-phosphate Cytidylyltransferase a(Gene symbol: Pcyt1a)
Molecule annotation
Chemicals (10 molecules)
1
2
2
1
3
7
* Click molecule labels to explore molecular sequence information.

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