Pyridoxine 5'-phosphate synthase

File:PdxJ mechanism.png removal.]]

In the first step of this condensation reaction, the amine group of HAP forms a Schiff base with the ketone group of DXP. The hydroxyl group on C4 of DXP is eliminated, forming an enol. The enol eliminates the phosphate derived from DXP, and water is added to the resulting double bond to reform the enol. This enol then attacks the HAP ketone group to close the ring and the resulting hydroxyl group is eliminated to form a double bond. A deprotonation causes the ring to aromatize, completing the synthesis of pyridoxine-5'-phosphate.

3-hydroxy-1-aminoacetone phosphate is unstable, so the reaction mechanism cannot be confirmed directly. Nonetheless, ¹⁴C and ¹⁸O isotopic labeling experiments,{{cite journal | vauthors = Cane DE, Du S, Robinson J | date = 1999 | title = Biosynthesis of vitamin B6: Enzymatic conversion of 1-deoxy-D-xylulose-5-phosphate to pyridoxol phosphate | journal = J. Am. Chem. Soc. | volume = 121 | pages = 7722–23 | doi = 10.1021/ja9914947 | issue = 33 }}{{cite journal | vauthors = Cane DE, Du S, Spenser ID | date = 2000 | title = Biosynthesis of vitamin B6: Origin of the oxygen atoms of pyridoxol phosphate | journal = J. Am. Chem. Soc. | volume = 122 | pages = 4213–14 | doi = 10.1021/ja000224h | issue = 17 }} as well as structural studies,{{cite journal | vauthors = Franco MG, Laber B, Huber R, Clausen T | title = Structural basis for the function of pyridoxine 5'-phosphate synthase | journal = Structure | volume = 9 | issue = 3 | pages = 245–53 | date = March 2001 | pmid = 11286891 | doi = 10.1016/S0969-2126(01)00584-6 | doi-access = free }} support the mechanism shown here. A glutamate residue, Glu72, is positioned ideally to perform most of the acid-base catalysis required in this mechanism, with histidine residues His45 and His193 appearing to play roles as well.

Pyridoxine-5'-phosphate synthase, or pdxJ, is a TIM barrel protein, although it exhibits some departures from this motif. Most significantly, the central tunnel of pdxJ is hydrophilic in contrast to the hydrophobic central tunnel observed in most TIM barrel proteins, and pdxJ has three extra alpha helices compared to the classical TIM fold.{{cite journal | vauthors = Garrido-Franco M | title = Pyridoxine 5'-phosphate synthase: de novo synthesis of vitamin B6 and beyond | journal = Biochimica et Biophysica Acta | volume = 1647 | issue = 1–2 | pages = 92–7 | date = April 2003 | pmid = 12686115 | doi = 10.1016/s1570-9639(03)00065-7 }} These three extra helices are important for mediating inter-subunit contacts in the assembled octamer.{{cite journal | vauthors = Fitzpatrick TB, Amrhein N, Kappes B, Macheroux P, Tews I, Raschle T | title = Two independent routes of de novo vitamin B6 biosynthesis: not that different after all | journal = The Biochemical Journal | volume = 407 | issue = 1 | pages = 1–13 | date = October 2007 | pmid = 17822383 | doi = 10.1042/bj20070765 | pmc = 2267407 }} However, there are also important similarities in function: like many TIM barrel proteins, pdxJ binds its substrates primarily by their phosphate moieties, and the phosphate-binding site responsible for binding to HAP and pyridoxine 5'-phosphate is a conserved motif found in many TIM barrel proteins.{{cite journal | vauthors = Nagano N, Orengo CA, Thornton JM | title = One fold with many functions: the evolutionary relationships between TIM barrel families based on their sequences, structures and functions | journal = Journal of Molecular Biology | volume = 321 | issue = 5 | pages = 741–65 | date = August 2002 | pmid = 12206759 | doi = 10.1016/S0022-2836(02)00649-6 }} The fact that pdxJ binds substrates through their phosphate groups explains a previously discovered specificity for the substrates over their respective non-phosphorylated alcohols.{{cite journal | vauthors = Laber B, Maurer W, Scharf S, Stepusin K, Schmidt FS | title = Vitamin B6 biosynthesis: formation of pyridoxine 5'-phosphate from 4-(phosphohydroxy)-L-threonine and 1-deoxy-D-xylulose-5-phosphate by PdxA and PdxJ protein | journal = FEBS Letters | volume = 449 | issue = 1 | pages = 45–8 | date = April 1999 | pmid = 10225425 | doi = 10.1016/S0014-5793(99)00393-2 | s2cid = 33542088 | doi-access = free }}

pdxJ exhibits several different conformations, depending on the substrates or substrate analogs bound. The first state, exhibited when pdxJ has either pyridoxine-5'-phosphate or no substrates bound, is classified as the "open" conformation. This conformation is characterized by an active site freely accessible by solvent. In contrast, when DXP and an HAP analog are bound, loop 4 of the protein folds over the active site, preventing the escape of reaction intermediates or undesirable side reactions. Binding of phosphate alone is not capable of causing a transition between the open and closed states. A third, "partially open" intermediate has also been reported upon binding of DXP alone.{{cite journal |vauthors=Yeh JI, Du S, Pohl E, Cane DE | date = 2002 | title = Multistate binding in pyridoxine 5′-phosphate synthase: 1.96 Å crystal structure in complex with 1-deoxy-D-xylulose phosphate | journal = Biochemistry | volume = 41 | pages = 11649–57 | issue = 39| doi=10.1021/bi026292t| pmid = 12269807 }}

pdxJ assembles as an octamer under biological conditions.{{cite journal |vauthors=Garrido-Franco M, Huber R, Schmidt FS, et al | date = 2000 | title = Crystallization and preliminary X-ray crystallographic analysis of PdxJ, the pyridoxine 50-phosphate synthesizing enzyme | journal = Acta Crystallographica | volume = 56 | issue = 8 | pages = 1045–48 | doi=10.1107/S0907444900007368| pmid = 10944349 | doi-access = }} This octamer can be thought of as a tetramer of dimers, and it is likely that the dimer is the active unit of the protein. In each dimer, an arginine residue Arg20 forms part of the active site in the other monomer, where it helps bind both phosphate groups.

This enzyme belongs to the family of transferases, specifically those transferring nitrogenous groups transferring other nitrogenous groups.

The systematic name of this enzyme class is 1-deoxy-D-xylulose-5-phosphate:3-amino-2-oxopropyl phosphate 3-amino-2-oxopropyltransferase (phosphate-hydrolysing; cyclizing). Other names in common use include pyridoxine 5-phosphate phospho lyase, PNP synthase, and PdxJ.

This enzyme participates in vitamin B₆ metabolism. pdxJ plays a role in the DXP-dependent pathway of pyridoxal phosphate. The DXP-dependent pathway is found predominantly in Gammaproteobacteria and some Alphaproteobacteria.{{cite journal | vauthors = Mittenhuber G | title = Phylogenetic analyses and comparative genomics of vitamin B6 (pyridoxine) and pyridoxal phosphate biosynthesis pathways | journal = Journal of Molecular Microbiology and Biotechnology | volume = 3 | issue = 1 | pages = 1–20 | date = January 2001 | pmid = 11200221 }} Because of this distribution, pdxJ has been identified as a potential drug target for antibiotics. This identification seems to have validity, as other approaches have also identified pdxJ as a good target for drug development.{{cite journal | vauthors = Ahmad S, Raza S, et al | date = 2018 | title = From phylogeny to protein dynamics: A computational hierarchical quest for potent drug identification against an emerging enteropathogen "Yersinia enterocolitica". | journal = Journal of Molecular Liquids | volume = 265 | pages = 372–89 | doi = 10.1016/j.molliq.2018.06.013 | s2cid = 90481778 }} However, there may be limits to this approach as pdxJ is not found in obligate parasites. pdxJ and more generally vitamin B₆ metabolism in the microbiome have also been shown to alter the effects of certain compounds on animal hosts.{{cite journal | vauthors = Scott TA, Quintaneiro LM, Norvaisas P, Lui PP, Wilson MP, Leung KY, Herrera-Dominguez L, Sudiwala S, Pessia A, Clayton PT, Bryson K, Velagapudi V, Mills PB, Typas A, Greene ND, Cabreiro F | display-authors = 6 | title = Host-Microbe Co-metabolism Dictates Cancer Drug Efficacy in C. elegans | journal = Cell | volume = 169 | issue = 3 | pages = 442–456.e18 | date = April 2017 | pmid = 28431245 | pmc = 5406385 | doi = 10.1016/j.cell.2017.03.040}}

{{reflist|30em}}

{{Nitrogenous transferases}}

{{Enzymes}}

{{Portal bar|Biology|border=no}}

Category:EC 2.6.99

Category:Enzymes of unknown structure