Substrate specificity of choline kinase

Clary, Greg L.; Tsai, Cheu-Fen; Guynn, Robert W.

doi:10.1016/0003-9861(87)90097-x

Cited by 32 publications

(9 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Nectrisine production by fungal necB disruptants and by recombinant E. coli without necB expression might be assisted by other enzymes catalyzing the NecB reaction with a variety of substrate specificities, such as choline kinases, most of which were shown to utilize both choline and ethanolamine as substrates (36)(37)(38). The existence of semiessential genes like necB in biosynthetic gene clusters has been reported previously; the aminotransferase gene tdiD was semiessential for bis-indolyquinone biosynthesis in Aspergillus nidulans (39).…”

Section: Discussionmentioning

confidence: 78%

Nectrisine Biosynthesis Genes in Thelonectria discophora SANK 18292: Identification and Functional Analysis

Miyauchi

Ono

Ohnuki

et al. 2016

Appl Environ Microbiol

View full text Add to dashboard Cite

The fungus Thelonectria discophora SANK 18292 produces the iminosugar nectrisine, which has a nitrogen-containing heterocyclic 5-membered ring and acts as a glycosidase inhibitor. In our previous study, an oxidase (designated NecC) that converts 4-amino-4-deoxyarabinitol to nectrisine was purified from T. discophora cultures. However, the genes required for nectrisine biosynthesis remained unclear. In this study, the nectrisine biosynthetic gene cluster in T. discophora was identified from the contiguous genome sequence around the necC gene. Gene disruption and complementation studies and heterologous expression of the gene showed that necA, necB, and necC could be involved in nectrisine biosynthesis, during which amination, dephosphorylation, and oxidation occur. It was also demonstrated that nectrisine could be produced by recombinant Escherichia coli coexpressing the necA, necB, and necC genes. These findings provide the foundation to develop a bacterial production system for nectrisine or its intermediates through genetic engineering. IMPORTANCEIminosugars might have great therapeutic potential for treatment of many diseases. However, information on the genes for their biosynthesis is limited. In this study, we report the identification of genes required for biosynthesis of the iminosugar nectrisine in Thelonectria discophora SANK 18292, which was verified by disruption, complementation, and heterologous expression of the genes involved. We also demonstrate heterologous production of nectrisine by recombinant E. coli, toward developing an efficient production system for nectrisine or its intermediates through genetic engineering.

show abstract

Section: Discussionmentioning

confidence: 78%

Nectrisine Biosynthesis Genes in Thelonectria discophora SANK 18292: Identification and Functional Analysis

Miyauchi

Ono

Ohnuki

et al. 2016

Appl Environ Microbiol

View full text Add to dashboard Cite

show abstract

“…TbC/EK2 was able to accept analogues modified at the amino group as substrates of the reaction; in fact, secondary and tertiary amino modifications were better than choline. Notably, of the compounds tested, ethanolamine was the poorest acceptor substrate (Table 2); these findings are in contrast with those with CK from S. cerevisiae [24], where N-methylethanolamine and N,N-dimethylethanolamine were as poor substrates as ethanolamine. Compounds with methyl and ethyl alterations on C-2 are good substrates only in the (S) configuration; R(−)2aminobutan-1-ol and R(−)2-aminopropan-1-ol are much poorer substrates, as well as 2-amino-2-methylpropan-1-ol, which bears two methyl groups at C-2 (Table 3).…”

Section: Acceptor Specificity Of Tbek1 and Tbc/ek2mentioning

confidence: 75%

“…Previously characterized C/EKs from various sources [15,[21][22][23][24][25] have exhibited different specificity towards the two substrates, choline and ethanolamine. It was therefore necessary to test the activity of the purified proteins in order to ascertain whether they were able to catalyse the phosphorylation of both choline and ethanolamine or if they showed selectivity to each branch of the Kennedy pathway.…”

Section: Kinetic Analysis Of Tbc/ek1 and Tbc/ek2mentioning

confidence: 99%

Biochemical characterization of the initial steps of the Kennedy pathway in Trypanosoma brucei: the ethanolamine and choline kinases

Gibellini¹,

Hunter²,

Smith³

2009

Biochemical Journal

View full text Add to dashboard Cite

Ethanolamine and choline are major components of the trypanosome membrane phospholipids, in the form of GPEtn (glycerophosphoethanolamine) and GPCho (glycerophosphocholine). Ethanolamine is also found as an integral component of the GPI (glycosylphosphatidylinositol) anchor that is required for membrane attachment of cell-surface proteins, most notably the variant-surface glycoproteins. The de novo synthesis of GPEtn and GPCho starts with the generation of phosphoethanolamine and phosphocholine by ethanolamine and choline kinases via the Kennedy pathway. Database mining revealed two putative C/EKs (choline/ethanolamine kinases) in the Trypanosoma brucei genome, which were cloned, overexpressed, purified and characterized. TbEK1 (T. brucei ethanolamine kinase 1) was shown to be catalytically active as an ethanolamine-specific kinase, i.e. it had no choline kinase activity. The K m values for ethanolamine and ATP were found to be 18.4 + − 0.9 and 219 + − 29 μM respectively. TbC/EK2 (T. brucei choline/ethanolamine kinase 2), on the other hand, was found to be able to phosphorylate both ethanolamine and choline, even though choline was the preferred substrate, with a K m 80 times lower than that of ethanolamine. The K m values for choline, ethanolamine and ATP were 31.4 + − 2.6 μM, 2.56 + − 0.31 mM and 20.6 + − 1.96 μM respectively. Further substrate specificity analysis revealed that both TbEK1 and TbC/EK2 were able to tolerate various modifications at the amino group, with the exception of a quaternary amine for TbEK1 (choline) and a primary amine for TbC/EK2 (ethanolamine). Both enzymes recognized analogues with substituents on C-2, but substitutions on C-1 and elongations of the carbon chain were not well tolerated.

show abstract

“…A small increase in the size of the quaternary ammonium head would sharply lower affinity for the transporter as large cholines failed to penetrate the cells. Intriguingly, another study (19) found that none of the choline analogs tested were better substrates than the simplest (native) choline for choline kinase with the exception of ethylcholine (Figure 2). A preclinical study confirmed this observation, in which the uptake of the native [ 11 C]-chloine in HCC was comparable to that of [ 18 F]-fluoroethylcholine (20).…”

Section: Limitationsmentioning

confidence: 90%

[18F]-choline PET/CT as an imaging biomarker for primary liver cancers

Lee

2016

Transl. Cancer Res.

View full text Add to dashboard Cite

Substrate specificity of choline kinase

Cited by 32 publications

References 22 publications

Nectrisine Biosynthesis Genes in Thelonectria discophora SANK 18292: Identification and Functional Analysis

Nectrisine Biosynthesis Genes in Thelonectria discophora SANK 18292: Identification and Functional Analysis

Biochemical characterization of the initial steps of the Kennedy pathway in Trypanosoma brucei: the ethanolamine and choline kinases

[18F]-choline PET/CT as an imaging biomarker for primary liver cancers

Contact Info

Product

Resources

About