2010 Vol. 1, No. 5

News and views
Repurposing an old anti-fungal drug as a Hedgehog inhibitor
Camilla Giambelli, Dennis Liang Fei, Huaizhi Wang, David J. Robbins
2010, 1(5): 417-418. doi: 10.1007/s13238-010-0063-5
Witness the prosperity: 25 years of life science revival in Tsinghua University
Zhirong Sun
2010, 1(5): 422-424. doi: 10.1007/s13238-010-0062-6
Return and reformation after a century-from 1918 to 2009 H1N1
Xiaoxue Zhang
2010, 1(5): 425-426. doi: 10.1007/s13238-010-0051-9
How to make a minimal genome for synthetic minimal cell
Liu-yan Zhang, Su-hua Chang, Jing Wang
2010, 1(5): 427-434. doi: 10.1007/s13238-010-0064-4
As a key focus of synthetic biology, building a minimal artificial cell has given rise to many discussions. A synthetic minimal cell will provide an appropriate chassis to integrate functional synthetic parts, devices and systems with functions that cannot generally be found in nature. The design and construction of a functional minimal genome is a key step while building such a cell/chassis since all the cell functions can be traced back to the genome. Kinds of approaches, based on bioinformatics and molecular biology, have been developed and proceeded to derive essential genes and minimal gene sets for the synthetic minimal genome. Experiments about streamlining genomes of model bacteria revealed genome reduction led to unanticipated beneficial properties, such as high electroporation efficiency and accurate propagation of recombinant genes and plasmids that were unstable in other strains. Recent achievements in chemical synthesis technology for large DNA segments together with the rapid development of the wholegenome sequencing, have transferred synthesis of genes to assembly of the whole genomes based on oligonucleotides, and thus created strong preconditions for synthesis of artificial minimal genome. Here in this article, we review briefly the history and current state of research in this field and summarize the main methods for making a minimal genome. We also discuss the impacts of minimized genome on metabolism and regulation of artificial cell.
Protein targets for structure-based antiMycobacterium tuberculosis drug discovery
Zhiyong Lou, Xiaoxue Zhang
2010, 1(5): 435-442. doi: 10.1007/s13238-010-0057-3
Mycobacterium tuberculosis, which belongs to the genus Mycobacterium, is the pathogenic agent for most tuberculosis (TB). As TB remains one of the most rampant infectious diseases, causing morbidity and death with emergence of multi-drug-resistant and extensively-drugresistant forms, it is urgent to identify new drugs with novel targets to ensure future therapeutic success. In this regards, the structural genomics of M. tuberculosis provides important information to identify potential targets, perform biochemical assays, determine crystal structures in complex with potential inhibitor(s), reveal the key sites/residues for biological activity, and thus validate drug targets and discover novel drugs. In this review, we will discuss the recent progress on novel targets for structure-based anti-M. tuberculosis drug discovery.
The structural basis for deadenylation by the CCR4-NOT complex
Mark Bartlam, Tadashi Yamamoto
2010, 1(5): 443-452. doi: 10.1007/s13238-010-0060-8
The CCR4-NOT complex is a highly conserved, multifunctional machinery controlling mRNA metabolism. Its components have been implicated in several aspects of mRNA and protein expression, including transcription initiation, elongation, mRNA degradation, ubiquitination, and protein modification. In this review, we will focus on the role of the CCR4-NOT complex in mRNA degradation. The complex contains two types of deadenylase enzymes, one belonging to the DEDD-type family and one belonging to the EEP-type family, which shorten the poly(A) tails of mRNA. We will review the present state of structure-function analyses into the CCR4-NOT deadenylases and summarize current understanding of their roles in mRNA degradation. We will also review structural and functional work on the Tob/BTG family of proteins, which are known to interact with the CCR4-NOT complex and which have been reported to suppress deadenylase activity in vitro.
Crystal structure of a novel non-Pfam protein PF2046 solved using low resolution B-factor sharpening and multi-crystal averaging methods
Jing Su, Yang Li, Neil Shaw, Weihong Zhou, Min Zhang, Hao Xu, Bi-Cheng Wang, Zhi-Jie Liu
2010, 1(5): 453-458. doi: 10.1007/s13238-010-0045-7
Sometimes crystals cannot diffract X-rays beyond 3.0 Å resolution due to the intrinsic flexibility associated with the protein. Low resolution diffraction data not only pose a challenge to structure determination, but also hamper interpretation of mechanistic details. Crystals of a 25.6 kDa non-Pfam, hypothetical protein, PF2046, diffracted X-rays to 3.38 Å resolution. A combination of SeMet derived heavy atom positions with multiple cycles of B-factor sharpening, multi-crystal averaging, restrained refinement followed by manual inspection of electron density and model building resulted in a final model with a R value of 23.5 (Rfree=24.7). The asymmetric unit was large and consisted of six molecules arranged as a homodimer of trimers. Analysis of the structure revealed the presence of a RNA binding domain suggesting a role for PF2046 in the processing of nucleic acids.
Research articles
Crystal structure of the swine-origin A (H1N1)-2009 influenza A virus hemagglutinin (HA) reveals similar antigenicity to that of the 1918 pandemic virus
Wei Zhang, Jianxun Qi, Yi Shi, Qing Li, Feng Gao, Yeping Sun, Xishan Lu, Qiong Lu, Christopher J. Vavricka, Di Liu, Jinghua Yan, George F. Gao
2010, 1(5): 459-467. doi: 10.1007/s13238-010-0059-1
Influenza virus is the causative agent of the seasonal and occasional pandemic flu. The current H1N1 influenza pandemic, announced by the WHO in June 2009, is highly contagious and responsible for global economic losses and fatalities. Although the H1N1 gene segments have three origins in terms of host species, the virus has been named swine-origin influenza virus (S-OIV) due to a predominant swine origin. 2009 S-OIV has been shown to highly resemble the 1918 pandemic virus in many aspects. Hemagglutinin is responsible for the host range and receptor binding of the virus and is therefore a primary indicator for the potential of infection. Primary sequence analysis of the 2009 S-OIV hemagglutinin (HA) reveals its closest relationship to that of the 1918 pandemic influenza virus, however, analysis at the structural level is necessary to critically assess the functional significance. In this report, we report the crystal structure of soluble hemagglutinin H1 (09H1) at 2.9 Å, illustrating that the 09H1 is very similar to the 1918 pandemic HA (18H1) in overall structure and the structural modules, including the five defined antiboby (Ab)-binding epitopes. Our results provide an explanation as to why sera from the survivors of the 1918 pandemics can neutralize the 2009 S-OIV, and people born around the 1918 are resistant to the current pandemic, yet younger generations are more susceptible to the 2009 pandemic.
Beclin 1 cleavage by caspase-3 inactivates autophagy and promotes apoptosis
Yushan Zhu, Lixia Zhao, Lei Liu, Ping Gao, Weili Tian, Xiaohui Wang, Haijing Jin, Haidong Xu, Quan Chen
2010, 1(5): 468-477. doi: 10.1007/s13238-010-0048-4
Autophagy and apoptosis are both highly regulated biological processes that play essential roles in tissue homeostasis, development and diseases. Autophagy is also described as a mechanism of death pathways, however, the precise mechanism of how autophagy links to cell death remains to be fully understood. Beclin 1 is a dual regulator for both autophagy and apoptosis. In this study we found that Beclin 1 was a substrate of caspase-3 with two cleavage sites at positions 124 and 149, respectively. Furthermore, the autophagosome formation occurred, followed by the appearance of morphological hallmarks of apoptosis after staurosporine treatment. The cleavage products of Beclin 1 reduced autophagy and promoted apoptosis in HeLa cells and the cells in which Beclin 1 was stably knocked down by specific shRNA. In addition, the cleavage of Beclin 1 resulted in abrogating the interaction between Bcl-2 with Beclin 1, which could be blocked by z-VAD-fmk. Thus, our results suggest that the cleavage of Beclin 1 by caspase-3 may contribute to inactivate autophagy leading towards augmented apoptosis.
Drosophila RecQ5 is required for efficient SSA repair and suppression of LOH in vivo
Yixu Chen, Wen Dui, Zhongsheng Yu, Changqing Li, Jun Ma, Renjie Jiao
2010, 1(5): 478-490. doi: 10.1007/s13238-010-0058-2
RecQ5 in mammalian cells has been suggested to suppress inappropriate homologous recombination. However, the specific pathway(s) in which it is involved and the underlining mechanism(s) remain poorly understood. We took advantage of genetic tools in Drosophila to investigate how Drosophila RecQ5 (dRecQ5) functions in vivo in homologous recombination-mediated double strand break (DSB) repair. We generated null alleles of dRecQ5 using the targeted recombination technique. The mutant animals are homozygous viable, but with growth retardation during development. The mutants are sensitive to both exogenous DSB-inducing treatment, such as gamma-irradiation, and endogenously induced double strand breaks (DSBs) by I-Sce I endonuclease. In the absence of dRecQ5, single strand annealing (SSA)-mediated DSB repair is compromised with compensatory increases in either inter-homologous gene conversion, or non-homologous end joining (NHEJ) when inter-chromosomal homologous sequence is unavailable. Loss of function of dRecQ5 also leads to genome instability in loss of heterozygosity (LOH) assays. Together, our data demonstrate that dRecQ5 functions in SSA-mediated DSB repair to achieve its full efficiency and in suppression of LOH in Drosophila.
Structures of EV71 RNA-dependent RNA polymerase in complex with substrate and analogue provide a drug target against the hand-foot-and-mouth disease pandemic in China
Yang Wu, Zhiyong Lou, Yi Miao, Yue Yu, Hui Dong, Wei Peng, Mark Bartlam, Xuemei Li, Zihe Rao
2010, 1(5): 491-500. doi: 10.1007/s13238-010-0061-7
Enterovirus 71 (EV71), one of the major causative agents for hand-foot-and-mouth disease (HFMD), has caused more than 100 deaths among Chinese children since March 2008. The EV71 genome encodes an RNAdependent RNA polymerase (RdRp), denoted 3Dpol, which is central for viral genome replication and is a key target for the discovery of specific antiviral therapeutics. Here we report the crystal structures of EV71 RdRp (3Dpol) and in complex with substrate guanosine-5'-triphosphate and analog 5-bromouridine-5'-triphosphate best to 2.4 Å resolution. The structure of EV71 RdRp (3Dpol) has a wider open thumb domain compared with the most closely related crystal structure of poliovirus RdRp. And the EV71 RdRp (3Dpol) complex with GTP or Br-UTP bounded shows two distinct movements of the polymerase by substrate or analogue binding. The model of the complex with the template:primer derived by superimposition with foot-and-mouth disease virus (FMDV) 3D/RNA complex reveals the likely recognition and binding of template:primer RNA by the polymerase. These results together provide a molecular basis for EV71 RNA replication and reveal a potential target for anti-EV71 drug discovery.