CTP:Phosphocholine Cytidylyltransferase: Paving the Way from Gene to Membrane

Jackowski, Suzanne; Fagone, Paolo

doi:10.1074/jbc.r400031200

Cited by 97 publications

(99 citation statements)

References 123 publications

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“…This reaction is catalyzed by CTP:phosphocholine cytidylyltransferase (CCT 4 ; EC 2.7.7.15), an enzyme subject to many layers of regulation (1)(2)(3)(4). The ubiquitous and best studied isoform of mammalian CCT (CCT␣, 367 residues) has been described as having four domains (Fig.…”

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confidence: 99%

Crystal Structure of a Mammalian CTP: Phosphocholine Cytidylyltransferase Catalytic Domain Reveals Novel Active Site Residues within a Highly Conserved Nucleotidyltransferase Fold

Lee

Johnson

Ding

et al. 2009

Journal of Biological Chemistry

104

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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 CCT␣ spanning residues 1-236 (CCT236) lacks the regulatory domain and as a result displays constitutive activity. The 2.2-Å 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 ␤-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.A key rate-limiting step in the synthesis of phosphatidylcholine in animal cells is the formation of the headgroup donor, CDP-choline, by transfer of a cytidylyl group from CTP to phosphocholine. This reaction is catalyzed by CTP:phosphocholine cytidylyltransferase (CCT 4 ; EC 2.7.7.15), an enzyme subject to many layers of regulation (1-4). The ubiquitous and best studied isoform of mammalian CCT (CCT␣, 367 residues) has been described as having four domains (Fig. 1A). An N-terminal domain (ϳ75 residues) housing its nuclear localization signal (NLS) sequence is followed by an ϳ150-residue catalytic domain, an ϳ60-residue membrane binding domain (domain M), and an unstructured phosphorylated tail (ϳ50 residues) (2, 4). CCT functions as a homodimer (5).CCT activation requires transformation of the enzyme from a soluble form to a membrane lipid-bound form. When the full-length soluble CCT interacts with anionic membrane surfaces, domain M transforms from a mixture of structural elements into an amphipathic ␣-helix (6 -8). Domain M appears to act as an autoinhibitory device, whose interaction with phosphatidylcholine-deficient membranes releases an inhibitory constraint at the active site to enhance k cat by 2 orders of magnitude (9). The primary evidence for this model is the constitutive activity of a construct lacking domain M, CCT236 (9).To elucidate the mechanism whereby membrane binding activates this important re...

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confidence: 99%

Crystal Structure of a Mammalian CTP: Phosphocholine Cytidylyltransferase Catalytic Domain Reveals Novel Active Site Residues within a Highly Conserved Nucleotidyltransferase Fold

Lee

Johnson

Ding

et al. 2009

Journal of Biological Chemistry

104

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“…The major route for PtdCho production is the CDP-choline pathway (11), and the supply of CDP-choline is governed by the activity of CCT. Three CCT isoforms are expressed differentially in tissues, but CCT␣ is the dominant isoform that is expressed ubiquitously (12,13).…”

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Phospholipid Biosynthesis Program Underlying Membrane Expansion during B-lymphocyte Differentiation

Fagone

Sriburi

Ward-Chapman

et al. 2007

Journal of Biological Chemistry

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“…Choline is used for production of phosphatidylcholine (PC) and sphingomyelin in the cycle called cytidine -diphosphocholine (CDP-choline) pathway. Phosphatidylcholine and sphingomyelin are essential for proper functioning of cell membranes, lipoprotein metabolism and moreover for the regulation of cellular life cycle [3,4]. PC is synthesized through the CDP-choline pathway and via the phosphatidylethanolamine N-methyltransferase (PEMT) de novo pathway.…”

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confidence: 99%

Polymorphic variants of genes involved in choline pathway and the risk of intrauterine fetal death

Drews¹,

Różycka²,

Barlik³

et al. 2017

Ginekol Pol

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Objectives: Choline and folate metabolism disturbances may be involved in the occurrence of intrauterine fetal death (IUFD). The proper activity of this metabolism could be determined by genetic variants involved in choline pathway e.g. CHKA (gene encoding choline kinase α), PCYT1A (gene encoding CCTα) and CHDH (gene encoding choline dehydrogenase). Our study aimed at determining the genotype and allele frequencies of CHKA rs7928739, PCYT1A rs712012, PCYT1A rs7639752, CHDH rs893363 and CHDH rs2289205 polymorphisms in mothers with IUFD occurrence. Material and methods:The study involved 76 mothers with IUFD occurrence and 215 mothers of healthy children. Genetic analysis was performed with the use of PCR/RFLP method. Results:The frequency of genotypes and alleles of studied polymorphisms was similar in both groups. The study revealed no association of PCYT1A, CHKA and CHDH polymorphisms in analysed groups of women. While evaluating the co-existence of analysed polymorphisms statistically significant correlation was revealed. Co-existence of CHKA rs7928739 AC/CHDH rs2289205 AA genotypes was observed statistically more frequently in the study group than in the control group (p = 0,031). Conclusions:There is no correlation between single CHKA rs7928739, PCYT1A rs712012, PCYT1A rs7639752, CHDH rs893363 and CHDH rs2289205 polymorphisms and the incidence of intrauterine fetal death. However, revealed statistically significant difference between co-existence of CHKA rs7928739 AC/CHDH rs2289205 AA genotypes between study groups suggest the need of further analysis.

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CTP:Phosphocholine Cytidylyltransferase: Paving the Way from Gene to Membrane

Cited by 97 publications

References 123 publications

Crystal Structure of a Mammalian CTP: Phosphocholine Cytidylyltransferase Catalytic Domain Reveals Novel Active Site Residues within a Highly Conserved Nucleotidyltransferase Fold

Crystal Structure of a Mammalian CTP: Phosphocholine Cytidylyltransferase Catalytic Domain Reveals Novel Active Site Residues within a Highly Conserved Nucleotidyltransferase Fold

Phospholipid Biosynthesis Program Underlying Membrane Expansion during B-lymphocyte Differentiation

Polymorphic variants of genes involved in choline pathway and the risk of intrauterine fetal death

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