Exploiting Enzymatic Promiscuity to Engineer a Focused Library of Highly Selective Antifungal and Antiproliferative Aureothin Analogues

Abstract

Aureothin is a shikimate-polyketide hybrid metabolite from Streptomyces thioluteus with a rare nitroaryl moiety, a chiral tetrahydrofuran ring and an O-methylated pyrone ring. The antimicrobial and antitumor activities of aureothin have caught our interest in modulating its structure as well as its bioactivity profile. In an integrated approach using mutasynthesis, biotransformation and combinatorial biosynthesis, a defined library of aureothin analogues was generated. The promiscuity of the polyketide synthase assembly line towards different starter units and the plasticity of the pyrone and tetrahydrofuran ring formation were exploited. A selection of fifteen new aureothin analogues with modifications at the aryl residue, the pyrone ring, and the oxygenated backbone was produced on a preparative scale and fully characterized. Remarkably, various new aureothin derivatives are less cytotoxic than aureothin but have improved antiproliferative activities. Furthermore, we found that the THF ring is crucial for the remarkably selective activity of aureothin analogues against certain pathogenic fungi.

Enzyme promiscuity: using the dark side of enzyme specificity in white biotechnology

“…The synthetic strategy avoids longer multistep processes. Werneburg et al [62] have used the polyketide synthetase for preparative scale synthesis of 15 new aureothin (a shikimate-polyketide with antimicrobial and antitumor activities) analogs, many with less cytotoxicity but improved anti-proliferative action. It may be noted that engineered mutants of the organism were used.…”

Metabolic Engineering for the Production of Natural Products

“…Recent extension of the work has led to the identification of two hybrid indolecarbazoles, which are potent and selective inhibitors of JAK2 and Ikkb kinases (141), underlying the potential of this strategy. More recently, Hertweck and co-workers used an integrated approach combining mutasynthesis (starter unit feeding), combinatorial biosynthesis (exchanging tailoring O- methyltransferase), and biotransformation (oxidations) to generate a focused library of 15 aureothin analogs, in which a few have less cytotoxicity but improved antiproliferative activities (142). …”

Novel Biotransformation Process of Podophyllotoxin to 4 -Sulfur-Substituted Podophyllum Derivates with Anti-Tumor Activity by Penicillium purpurogenum Y.J. Tang

“…According to bioisosterism, the O-linked (ethers, esters) compounds could be less active in comparison to the S-linked congeners. So, the sulfur atom preferentially integrated with the large biological protein molecule with the hydrogen bond in tumor cell, and the sulfur-substituted is an effective pathway to improve anti-tumor activity and reduce toxic side-effects of the *Address correspondence to this author at the Key Laboratory of Fermentation Engineering (Ministry of Education), Hubei University of Technology, Wuhan 430068, China; Tel/Fax (+86-27)-8801.5108; E-mail: yajietang@yahoo.com.cn leading compounds [12][13][14]. On the other hand, by comparing carbon atom with oxygen atom in the hydroxyl group of PTOX, the carbon atom with positive charge is more reactive.…”

“…R f =0.55 (CHCl 3 /Acetone 2:1); for 1 H (300 MHz, CDCl 3 , 25 , TMS) and13 C (75 MHz, CDCl 3 , 25 , TMS (75 MHz) NMR spectra, see Table 1. 1 H NMR: =8.19 (s, 1H; CH), 7.02 (s, 1H; Ar-H), 6.47 (s, 1H; Ar-H), 6.31 (s, 2H; Ar-H), 5.98 (d, 2 J(13,13)=5.1 Hz, 2H; OCH 2 O), 5.38 (s, 1H; CH), 4.60 (s, 1H; CH), 4.35 (s, 2 J(11,11)=5.7 Hz, 1H; CH 2 ), 3.99 (s, 1H; CH 2 ), 3.81 (t, 2 J(3',3')=7.2Hz, 9H; OCH 3 ), 3.28 ppm (s, 2H, CH); 13 C NMR: =174.8 (COOCH 2 ), 158.6 (CNH), 152.7 (CHN, COCH 3 ), 148.6 (CCH), 147.7 (CCH), 137.3 (COCH 3 ), 135.7 (CCH), 132.5 (CCH), 127.6 (CCH), 110.4 (CH), 110.1 (CH), 108.5 (CH), 101.9 (OCH 2 O), 70.8(CH 2 ), 61.0 (OCH 3 ), 56.5 (OCH 3 ), 49.1 (CH), 43.9(CH), 42.4 (CH), 37.489 ppm (CH); IR (Nujol): ˜=1093.7 cm -1 (C[sbond]S); UV: max ( )=221; EI MS: m/z:499 [M + +2H], 521 [M + +Na+H], 537 [M + +K+H]; elemental analysis calcd (%) for C 24 H 23 O 7 N 3 S 1 : C 57.94, H 4.66, O 22.51, N 8.45, S 6.45; found C 55.88, H 4.66, O 22.76, N 7.90, S 5.10.…”