| Structural highlights
Function
CP51B_ASPFU Sterol 14-alpha demethylase; part of the third module of ergosterol biosynthesis pathway that includes the late steps of the pathway (PubMed:18191972, PubMed:26459890, PubMed:29439966, PubMed:9184358). Demethylates eburicol to yield 4,4,24-trimethyl ergosta-8,14,24(28)-trienol (PubMed:18191972, PubMed:26459890, PubMed:29439966, PubMed:9184358). The third module or late pathway involves the ergosterol synthesis itself through consecutive reactions that mainly occur in the endoplasmic reticulum (ER) membrane. Firstly, the squalene synthase erg9 catalyzes the condensation of 2 farnesyl pyrophosphate moieties to form squalene, which is the precursor of all steroids. Squalene synthase is crucial for balancing the incorporation of farnesyl diphosphate (FPP) into sterol and nonsterol isoprene synthesis. Secondly, squalene is converted into lanosterol by the consecutive action of the squalene epoxidase erg1 and the lanosterol synthase erg7. Then, the delta(24)-sterol C-methyltransferase erg6 methylates lanosterol at C-24 to produce eburicol. Eburicol is the substrate of the sterol 14-alpha demethylase encoded by cyp51A and cyp51B, to yield 4,4,24-trimethyl ergosta-8,14,24(28)-trienol. The C-14 reductase erg24 then reduces the C14=C15 double bond which leads to 4,4-dimethylfecosterol. A sequence of further demethylations at C-4, involving the C-4 demethylation complex containing the C-4 methylsterol oxidases erg25A or erg25B, the sterol-4-alpha-carboxylate 3-dehydrogenase erg26 and the 3-keto-steroid reductase erg27, leads to the production of fecosterol via 4-methylfecosterol. The C-8 sterol isomerase erg2 then catalyzes the reaction which results in unsaturation at C-7 in the B ring of sterols and thus converts fecosterol to episterol. The sterol-C5-desaturase erg3B then catalyzes the introduction of a C-5 double bond in the B ring to produce 5-dehydroepisterol. The 2 other sterol-C5-desaturases, erg3A and erg3C, seem to be less important in ergosterol biosynthesis. The C-22 sterol desaturase erg5 further converts 5-dehydroepisterol into ergosta-5,7,22,24(28)-tetraen-3beta-ol by forming the C-22(23) double bond in the sterol side chain. Finally, ergosta-5,7,22,24(28)-tetraen-3beta-ol is substrate of the C-24(28) sterol reductases erg4A and erg4B to produce ergosterol. Possible alternative sterol biosynthetic pathways might exist from fecosterol to ergosterol, depending on the activities of the erg3 isoforms (PubMed:16110826, PubMed:18191972) (Probable).[1] [2] [3] [4] [5] [6] As a target of azole drugs, plays a crucial role in azole susceptibility.[7] [8] [9] [10]
Publication Abstract from PubMed
Aspergillus fumigatus is the opportunistic fungal pathogen that predominantly affects the immunocompromised population and causes 600,000 deaths per year. The cytochrome P450 (CYP) 51 inhibitor voriconazole is currently the drug of choice, yet the treatment efficiency remains low, calling for rational development of more efficient agents. A. fumigatus has two CYP51 genes, CYP51A and CYP51B, which share 59% amino acid sequence identity. CYP51B is expressed constitutively, while gene CYP51A is reported to be inducible. We expressed, purified, and characterized A. fumigatus CYP51B, including determination of its substrate preferences, catalytic parameters, inhibition, and X-ray structure in complexes with voriconazole and the experimental inhibitor (R)-N-(1-(2,4-dichlorophenyl)-2-(1H-imidazol-1-yl)ethyl)-4-(5-phenyl-1,3,4-oxadia zol-2-yl)benzamide (VNI). The enzyme demethylated its natural substrate eburicol and the plant CYP51 substrate obtusifoliol at steady-state rates of 17 and 16 min-1, respectively, but did not metabolize lanosterol, and the topical antifungal drug miconazole was the strongest inhibitor that we identified. The X-ray crystal structures displayed high overall similarity of A. fumigatus CYP51B to CYP51 orthologs from other biological kingdoms but revealed phylum-specific differences relevant to enzyme catalysis and inhibition. The complex with voriconazole provides an explanation for the potency of this relatively small molecule, while the complex with VNI outlines a direction for further enhancement of the efficiency of this new inhibitory scaffold to treat humans afflicted with filamentous fungal infections.
Structure-functional Characterization of Cytochrome P450 Sterol 14alpha-Demethylase (CYP51B) from Aspergillus fumigatus and Molecular Basis for the Development of Antifungal Drugs.,Hargrove TY, Wawrzak Z, Lamb DC, Guengerich FP, Lepesheva GI J Biol Chem. 2015 Aug 12. pii: jbc.M115.677310. PMID:26269599[11]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
- ↑ Alcazar-Fuoli L, Mellado E, Garcia-Effron G, Lopez JF, Grimalt JO, Cuenca-Estrella JM, Rodriguez-Tudela JL. Ergosterol biosynthesis pathway in Aspergillus fumigatus. Steroids. 2008 Mar;73(3):339-47. PMID:18191972 doi:10.1016/j.steroids.2007.11.005
- ↑ Warrilow AG, Parker JE, Price CL, Nes WD, Kelly SL, Kelly DE. In Vitro Biochemical Study of CYP51-Mediated Azole Resistance in Aspergillus fumigatus. Antimicrob Agents Chemother. 2015 Dec;59(12):7771-8. PMID:26459890 doi:10.1128/AAC.01806-15
- ↑ Colley T, Sehra G, Chowdhary A, Alanio A, Kelly SL, Kizawa Y, Armstrong-James D, Fisher MC, Warrilow AGS, Parker JE, Kelly DE, Kimura G, Nishimoto Y, Sunose M, Onions S, Crepin D, Lagasse F, Crittall M, Shannon J, McConville M, King-Underwood J, Naylor A, Bretagne S, Murray J, Ito K, Strong P, Rapeport G. In Vitro and In Vivo Efficacy of a Novel and Long-Acting Fungicidal Azole, PC1244, on Aspergillus fumigatus Infection. Antimicrob Agents Chemother. 2018 Apr 26;62(5):e01941-17. PMID:29439966 doi:10.1128/AAC.01941-17
- ↑ Venkateswarlu K, Kelly SL. Stereoselective interaction of SCH 39304, a triazole, with sterol 14alpha-demethylase of Aspergillus fumigatus. J Antimicrob Chemother. 1997 May;39(5):597-601. PMID:9184358 doi:10.1093/jac/39.5.597
- ↑ Ferreira ME, Colombo AL, Paulsen I, Ren Q, Wortman J, Huang J, Goldman MH, Goldman GH. The ergosterol biosynthesis pathway, transporter genes, and azole resistance in Aspergillus fumigatus. Med Mycol. 2005 May;43 Suppl 1:S313-9. PMID:16110826 doi:10.1080/13693780400029114
- ↑ Alcazar-Fuoli L, Mellado E, Garcia-Effron G, Lopez JF, Grimalt JO, Cuenca-Estrella JM, Rodriguez-Tudela JL. Ergosterol biosynthesis pathway in Aspergillus fumigatus. Steroids. 2008 Mar;73(3):339-47. PMID:18191972 doi:10.1016/j.steroids.2007.11.005
- ↑ Mann PA, Parmegiani RM, Wei SQ, Mendrick CA, Li X, Loebenberg D, DiDomenico B, Hare RS, Walker SS, McNicholas PM. Mutations in Aspergillus fumigatus resulting in reduced susceptibility to posaconazole appear to be restricted to a single amino acid in the cytochrome P450 14alpha-demethylase. Antimicrob Agents Chemother. 2003 Feb;47(2):577-81. PMID:12543662 doi:10.1128/AAC.47.2.577-581.2003
- ↑ Hargrove TY, Wawrzak Z, Lamb DC, Guengerich FP, Lepesheva GI. Structure-functional Characterization of Cytochrome P450 Sterol 14alpha-Demethylase (CYP51B) from Aspergillus fumigatus and Molecular Basis for the Development of Antifungal Drugs. J Biol Chem. 2015 Aug 12. pii: jbc.M115.677310. PMID:26269599 doi:http://dx.doi.org/10.1074/jbc.M115.677310
- ↑ Hargrove TY, Garvey EP, Hoekstra WJ, Yates CM, Wawrzak Z, Rachakonda G, Villalta F, Lepesheva GI. Crystal structure of the new investigational drug candidate VT-1598 in complex with Aspergillus fumigatus sterol 14alpha-demethylase provides insights into its broad-spectrum antifungal activity. Antimicrob Agents Chemother. 2017 May 1. pii: AAC.00570-17. doi:, 10.1128/AAC.00570-17. PMID:28461309 doi:http://dx.doi.org/10.1128/AAC.00570-17
- ↑ Friggeri L, Hargrove TY, Wawrzak Z, Blobaum AL, Rachakonda G, Lindsley CW, Villalta F, Nes WD, Botta M, Guengerich FP, Lepesheva GI. Sterol 14alpha-Demethylase Structure-Based Design of VNI (( R)- N-(1-(2,4-Dichlorophenyl)-2-(1 H-imidazol-1-yl)ethyl)-4-(5-phenyl-1,3,4-oxadiazol-2-yl)benzamide)) Derivatives To Target Fungal Infections: Synthesis, Biological Evaluation, and Crystallographic Analysis. J Med Chem. 2018 Jun 25. doi: 10.1021/acs.jmedchem.8b00641. PMID:29894182 doi:http://dx.doi.org/10.1021/acs.jmedchem.8b00641
- ↑ Hargrove TY, Wawrzak Z, Lamb DC, Guengerich FP, Lepesheva GI. Structure-functional Characterization of Cytochrome P450 Sterol 14alpha-Demethylase (CYP51B) from Aspergillus fumigatus and Molecular Basis for the Development of Antifungal Drugs. J Biol Chem. 2015 Aug 12. pii: jbc.M115.677310. PMID:26269599 doi:http://dx.doi.org/10.1074/jbc.M115.677310
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