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Deoxyribose-phosphate aldolase

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deoxyribose-phosphate aldolase
Identifiers
EC no.4.1.2.4
CAS no.9026-97-5
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The enzyme deoxyribose-phosphate aldolase (EC 4.1.2.4) catalyzes the reversible chemical reaction

2-deoxy-D-ribose 5-phosphate D-glyceraldehyde 3-phosphate + acetaldehyde

This enzyme belongs to the family of lyases, specifically the aldehyde-lyases, which cleave carbon-carbon bonds. The systematic name of this enzyme class is 2-deoxy-D-ribose-5-phosphate acetaldehyde-lyase (D-glyceraldehyde-3-phosphate-forming). Other names in common use include phosphodeoxyriboaldolase, deoxyriboaldolase, deoxyribose-5-phosphate aldolase, 2-deoxyribose-5-phosphate aldolase, and 2-deoxy-D-ribose-5-phosphate acetaldehyde-lyase.

Enzyme Mechanism

Mechanism of DERA catalysis. First, substrate is shown, followed by stabilizing interactions at the active site. Finally, key lysine residues and the carbinolamine intermediate are shown. Based on PDB 1JCL

Amongst aldolases, DERA is unique as it is the only enzyme to afford two aldehydes as products.[1] Crystallography shows that the enzyme is a Class I aldolase, so the mechanism proceeds via the formation of a Schiff base with Lys167 at the active site. A nearby residue, Lys201, is critical to reaction by increasing the acidity of protonated Lys167, so Schiff base formation can occur more readily.[2]

As equilibrium of the reaction as written lies on the side of reactant, DERA can also used to catalyze the backward aldol reaction. The enzyme has been found to exhibit some promiscuity by accepting various carbonyl compounds as substrates: acetaldehyde can be replaced with other small aldehydes or acetone; and a variety of aldehydes can be used in place of D-glyceraldehyde 3-phosphate. However, due to the spatial arrangement of stabilizing interactions of the electrophilic aldehyde at the active site, the aldol reaction is stereospecific and gives the (S)-configuration at the reactive carbon. Molecular modeling of the active site showed a hydrophilic pocket formed by Thr170 and Lys172 to stabilize C2-hydroxy group of D-glyceraldehyde 3-phosphate, while the C2-hydrogen atom is stabilized in a hydrophobic pocket. When a racemic mixture of glyceraldehyde 3-phosphate is used as the substrate, only the D-isomer reacted.[3]

Enzyme Structure

The DERA monomer contains a TIM α/β barrel fold, consistent with other Class I aldolases.[2] The structure of DERAs across many organisms: DERAs from Escherichia coli and Aeropyrum pernix shares 37.7% sequence identity with DERA from Thermus thermophilus HB8.[4] The reaction mechanism is also conserved between DERAs.

In solution, DERAs are found in homodimers or homotetramers. The oligomeric nature of the enzyme does not contribute to enzymatic activity, but serves to increase thermal stability through hydrophobic interactions and hydrogen bonding between interfacial residues.[5]

As of late 2007, 10 structures have been solved for this class of enzymes, with PDB accession codes 1JCJ, 1JCL, 1KTN, 1MZH, 1N7K, 1O0Y, 1P1X, 1UB3, 1VCV, and 2A4A.

Biological Function

DERA is part of the inducible deo operon in bacteria which allows for the conversion of exogenous deoxyribonucleosides for energy generation.[6] The products of DERA, glyceraldehyde-3-phosphate and acetaldehyde (subsequently converted to acetyl CoA) can enter the glycolysis and Kreb’s cycle pathways respectively.

In humans, DERA is mainly expressed in lungs, liver and colon and is necessary for the cellular stress response. After induction of oxidative stress or mitochondrial stress, DERA colocalizes with stress granules and associates with YBX1, a known stress granule protein. Cells with high DERA expression were able to utilize exogenous deoxyinosine to produce ATP when starved of glucose and incubated with mitochondrial uncoupler FCCP.[7]

Industrial Relevance

DERA used in islatravir biocatalysis. Bonds formed by DERA are highlighted in red.

DERA is being used in chemical syntheses as a tool for green, enantioselective aldol reactions. Formation of the deoxyribose skeleton from small molecules can facilitate the synthesis of nucleoside reverse transcriptase inhibitors.[8] For example, DERA was used in a mixture of five enzymes in the biocatalytic synthesis of islatravir.[9]

DERA used in Atorvastatin biocatalysis. The portion of Atorvastatin which was derived from the DERA catalyzed reaction is indicated.

DERA has also been used to perform tandem aldol reactions with three aldehyde substrates, with reaction equilibrium driven by the formation of the six-membered cyclic hemiacetal.[10] This intermediate has been used in the synthesis of statin drugs, such as atorvastatin,[11] rosuvastatin and mevastatin.[12]

Natural DERAs show low tolerance to high concentrations of acetaldehyde[13] due to the formation of the highly reactive crotonaldehyde intermediate that irreversibly inactivates the enzyme.[14] This features hampers the industrial applications of DERA as the concentration of acetaldehyde used will be limited. To overcome this, directed evolution has been used to improve the acetaldehyde tolerance of DERA to up to 400mM.[9]

References

  1. ^ Chambre, Domitille; Guérard-Hélaine, Christine; Darii, Ekaterina; Mariage, Aline; Petit, Jean-Louis; Salanoubat, Marcel; de Berardinis, Véronique; Lemaire, Marielle; Hélaine, Virgil (2019). "2-Deoxyribose-5-phosphate aldolase, a remarkably tolerant aldolase towards nucleophile substrates". Chemical Communications. 55 (52): 7498–7501. doi:10.1039/c9cc03361k. PMID 31187106.
  2. ^ a b Heine, Andreas; Luz, John G.; Wong, Chi-Huey; Wilson, Ian A. (October 2004). "Analysis of the Class I Aldolase Binding Site Architecture Based on the Crystal Structure of 2-Deoxyribose-5-phosphate Aldolase at 0.99Å Resolution". Journal of Molecular Biology. 343 (4): 1019–1034. doi:10.1016/j.jmb.2004.08.066. PMID 15476818.
  3. ^ Liu, Junjie; Wong, Chi-Huey (15 April 2002). "Aldolase-Catalyzed Asymmetric Synthesis of Novel Pyranose Synthons as a New Entry to Heterocycles and Epothilones". Angewandte Chemie International Edition. 41 (8): 1404–1407. doi:10.1002/1521-3773(20020415)41:8<1404::AID-ANIE1404>3.0.CO;2-G. PMID 19750780.
  4. ^ Lokanath, N. K.; Shiromizu, I.; Ohshima, N.; Nodake, Y.; Sugahara, M.; Yokoyama, S.; Kuramitsu, S.; Miyano, M.; Kunishima, N. (1 October 2004). "Structure of aldolase from Thermus thermophilus HB8 showing the contribution of oligomeric state to thermostability". Acta Crystallographica Section D. 60 (10): 1816–1823. doi:10.1107/S0907444904020190. ISSN 0907-4449. PMID 15388928.
  5. ^ Dick, Markus; Weiergräber, Oliver H.; Classen, Thomas; Bisterfeld, Carolin; Bramski, Julia; Gohlke, Holger; Pietruszka, Jörg (19 January 2016). "Trading off stability against activity in extremophilic aldolases". Scientific Reports. 6 (1): 17908. Bibcode:2016NatSR...617908D. doi:10.1038/srep17908. PMC 4725968. PMID 26783049.
  6. ^ Lomax, MS; Greenberg, GR (August 1968). "Characteristics of the deo operon: role in thymine utilization and sensitivity to deoxyribonucleosides". Journal of Bacteriology. 96 (2): 501–14. doi:10.1128/JB.96.2.501-514.1968. PMC 252324. PMID 4877128.
  7. ^ Salleron, Lisa; Magistrelli, Giovanni; Mary, Camille; Fischer, Nicolas; Bairoch, Amos; Lane, Lydie (December 2014). "DERA is the human deoxyribose phosphate aldolase and is involved in stress response". Biochimica et Biophysica Acta (BBA) - Molecular Cell Research. 1843 (12): 2913–2925. doi:10.1016/j.bbamcr.2014.09.007. PMID 25229427.
  8. ^ HORINOUCHI, Nobuyuki; OGAWA, Jun; KAWANO, Takako; SAKAI, Takafumi; SAITO, Kyota; MATSUMOTO, Seiichiro; SASAKI, Mie; MIKAMI, Yoichi; SHIMIZU, Sakayu (22 May 2014). "Efficient Production of 2-Deoxyribose 5-Phosphate from Glucose and Acetaldehyde by Coupling of the Alcoholic Fermentation System of Baker's Yeast and Deoxyriboaldolase-Expressing". Bioscience, Biotechnology, and Biochemistry. 70 (6): 1371–1378. doi:10.1271/bbb.50648. PMID 16794316. S2CID 22227871.
  9. ^ a b Huffman, Mark A.; Fryszkowska, Anna; Alvizo, Oscar; Borra-Garske, Margie; Campos, Kevin R.; Canada, Keith A.; Devine, Paul N.; Duan, Da; Forstater, Jacob H.; Grosser, Shane T.; Halsey, Holst M.; Hughes, Gregory J.; Jo, Junyong; Joyce, Leo A.; Kolev, Joshua N.; Liang, Jack; Maloney, Kevin M.; Mann, Benjamin F.; Marshall, Nicholas M.; McLaughlin, Mark; Moore, Jeffrey C.; Murphy, Grant S.; Nawrat, Christopher C.; Nazor, Jovana; Novick, Scott; Patel, Niki R.; Rodriguez-Granillo, Agustina; Robaire, Sandra A.; Sherer, Edward C.; Truppo, Matthew D.; Whittaker, Aaron M.; Verma, Deeptak; Xiao, Li; Xu, Yingju; Yang, Hao (5 December 2019). "Design of an in vitro biocatalytic cascade for the manufacture of islatravir". Science. 366 (6470): 1255–1259. Bibcode:2019Sci...366.1255H. doi:10.1126/science.aay8484. PMID 31806816. S2CID 208738388.
  10. ^ GIJSEN, H. J. M.; WONG, C.-H. (18 August 2010). "ChemInform Abstract: Unprecedented Asymmetric Aldol Reactions with Three Aldehyde Substrates Catalyzed by 2-Deoxyribose-5-phosphate Aldolase". ChemInform. 26 (15): no. doi:10.1002/chin.199515259. ISSN 0931-7597.
  11. ^ Jennewein, Stefan; Schürmann, Martin; Wolberg, Michael; Hilker, Iris; Luiten, Ruud; Wubbolts, Marcel; Mink, Daniel (May 2006). "Directed evolution of an industrial biocatalyst: 2-deoxy-D-ribose 5-phosphate aldolase". Biotechnology Journal. 1 (5): 537–548. doi:10.1002/biot.200600020. PMID 16892289.
  12. ^ Patel, Ramesh N. (April 2018). "Biocatalysis for synthesis of pharmaceuticals". Bioorganic & Medicinal Chemistry. 26 (7): 1252–1274. doi:10.1016/j.bmc.2017.05.023. PMID 28648492.
  13. ^ Haridas, Meera; Abdelraheem, Eman M. M.; Hanefeld, Ulf (3 October 2018). "2-Deoxy-d-ribose-5-phosphate aldolase (DERA): applications and modifications". Applied Microbiology and Biotechnology. 102 (23): 9959–9971. doi:10.1007/s00253-018-9392-8. PMC 6244999. PMID 30284013.
  14. ^ Dick, Markus; Hartmann, Rudolf; Weiergräber, Oliver H.; Bisterfeld, Carolin; Classen, Thomas; Schwarten, Melanie; Neudecker, Philipp; Willbold, Dieter; Pietruszka, Jörg (2016). "Mechanism-based inhibition of an aldolase at high concentrations of its natural substrate acetaldehyde: structural insights and protective strategies". Chemical Science. 7 (7): 4492–4502. doi:10.1039/c5sc04574f. PMC 6016325. PMID 30155096.
  • Hoffee PA (1968). "2-deoxyribose-5-phosphate aldolase of Salmonella typhimurium: purification and properties". Arch. Biochem. Biophys. 126 (3): 795–802. doi:10.1016/0003-9861(68)90473-6. PMID 4879701.
  • Jedziniak JA, Lionetti FJ (1970). "Purification and properties of deoxyriboaldolase from human erythrocytes". Biochim. Biophys. Acta. 212 (3): 478–87. doi:10.1016/0005-2744(70)90254-8. PMID 4989681.
  • Racker E (1952). "Enzymatic synthesis and breakdown of desoxyribose phosphate". J. Biol. Chem. 196 (1): 347–65. PMID 12980976.
  • Hoffee P, Rosen OM, Horecker BL (1965). "The mechanism of action of aldolases. VI. Crystallization of deoxyribose 5-phosphate aldolase and the number of active sites". J. Biol. Chem. 240: 1512–1516. PMID 14285485.
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