Jingjing Zhou, Chengbin Qiu, Sicong Li, Xiyao Cheng, Lei Zhang, Yao Chen, Yongqi Huang*, Yuhui Sun*, and Zhengding Su*
ChemistrySelect 2018, 3(39):33048-11053
Epub Date: 24 October 2018
The readthrough of premature termination codons (PTCs) is a promising strategy for curing PTC-causing diseases. In cancers, the p53 anti-tumor activity is often disabledby forming premature terminated p53 (p53PTC). Currently, there arelack of p53PTC-rescuing drugs. Herein we designed a feasible strategy for identifying p53PTC-rescuing compounds using protein biosynthesismachinery in E. coli cells and lung cancer H1299 cells both harboring p53PTC-GFP fusion protein expression cassettes. Rescued p53PTC enabled a GFP-tag for fluorescence spectroscopic assays. Our data revealed that the aminoglycoside G418 not only efficiently rescued p53PTC in H1299 cells, but also synergistically enhanced the efficacy of antitumor drug doxotubicin (DOX). Our work lends insighton accelerating the discovery of p53PTC-rescuing drugs for cancertherapy.
Weixin Tao, Anna Yang, Zixin Deng, and Yuhui Sun*
Frontiers in Microbiology 2018, 9:1660
Epub Date: 24 July 2018
Microbial natural products (NPs) especially of the Streptomyces genus have been regarded as an unparalleled resource for pharmaceutical drugs discovery. Moreover, recentprogress in sequencing technologies and computational resources further reinforces to identify numerous NP biosynthetic gene clusters (BGCs) from the genomes of Streptomyces. However, the majority of these BGCs are silent orpoorly expressed in native strains and remain to be activated and investigated, which relies heavily on efficient genome editing approaches. Accordingly, numerous strategies are developed, especially, the most recently developed, namely, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated (Cas) system reveals remarkable higher accuracy and efficiency for genome editing in various model organisms including the Streptomyces. In this mini review, we highlight the application of CRISPR/Cas9-based approaches in Streptomyces, focus on the editing of BGCs either in vivo or in vitro, as well as target cloning of large-sized BGCs and heterologous expression in a genetically manipulatable host, for discovery, characterization, reengineering, and production of potential pharmaceutical drugs.
Sicong Li#, Jun Zhang#, Yuanzhen Liu, Guo Sun, Zixin Deng, and Yuhui Sun*
ACS Chemical Biology 2018, 13(8):2203-2210
Epub Date: 19 June 2018
HygromycinB is an aminoglycoside antibiotic with a structurally distinctive orthoester linkage. Despite its long history of use in industry and in the laboratory, its biosynthesis remains poorly understood. We show here, by in-frame gene deletion in vivo and detailed enzyme characterization in vitro, that formation of the unique orthoester moiety is catalyzed by the α-ketoglutarat- and non-heme iron-dependent oxygenase HygX. In addition, we identify HygF as aglycosyl transferase adding UDP-hexose to 2-deoxystreptamine, HygM as a methyltransferase responsible for N-3 methylation, and HygK as an epimerase. These experimental results and bioinformatic analyses allow a detailed pathway for hygromycin B biosynthesis to be proposed, including the key oxidative cyclization reactions.
Yuan Li#, Wan Zhang#, Hui Zhang, Wenya Tian, Lian Wu, Shuwen Wang, Mengmeng Zheng, Jinru Zhang, Chenghai Sun, Zixin Deng, Yuhui Sun, Xudong Qu*, and Jiahai Zhou*
Angewandte Chemie International Edition 2018, 57(20):5823-5827
Epub Date: 14 April 2018
Polyketides are a large family of pharmaceutically important natural products, and the structural modification of their scaffolds is significant for drug development. Herein, we report high‐resolution X‐ray crystal structures of the broadly selective acyltransferase (AT) from the splenocin polyketide synthase (SpnD‐AT) in the apo form and in complex with benzylmalonyl and pentynylmalonyl extender unit mimics. These structures revealed the molecular basis for the stereoselectivity and substrate specificity of SpnD‐AT, and enabled the engineering of the industrially important Ery‐AT6 to broaden its substrate scope to include three new types of extender units.
Sicong Li#, Junhong Guo#, Anna Reva, Fanglu Huang, Binbin Xiong, Yuanzhen Liu, Zixin Deng, Peter F. Leadlay*, and YuhuiSun*
Proceedings of the National Academy of Sciences of the United States of America 2018, 115(6):1340-1345
Epub Date: 22 January 2018
Aminoglycosides remain a vital clinical asset. Gentamicin C complex in particular is remarkably potent in treating systemic Gram-negative infections, and semisynthetic gentamicins that combat pathogen resistance or show reduced toxicity remain attractive goals. We report here the roles of clustered genes and enzymes that define a methylation network in gentamicin biosynthesis and also identify a remote gene on the chromosome encoding the essential methyltransferase GenL, which is decisive for the proportions of the five major components present in the gentamicin C complex. This is an important step toward engineered fermentation to produce single components as valuable starting materials for semisynthesis of next-generation aminoglycoside antibiotics.