玫瑰杆菌类群细菌参与海洋有机硫DMSP去甲基化代谢关键酶DmdB和DmdC的结构与催化机制

项目来源

国家自然科学基金(NSFC)

项目主持人

王鹏

项目受资助机构

山东大学

项目编号

31800107

立项年度

2018

立项时间

未公开

项目级别

国家级

研究期限

未知 / 未知

受资助金额

26.00万元

学科

生命科学-微生物学-微生物与环境互作

学科代码

C-C01-C0106

基金类别

青年科学基金项目

关键词

海洋微生物 ; 生物地球化学循环 ; 玫瑰杆菌 ; 二甲基巯基丙酸内盐 ; 去甲基化代谢 ; 海洋微生物 ; 生物地球化学循环 ; 玫瑰杆菌 ; 二甲基巯基丙酸内盐 ; 去甲基化代谢

参与者

李春阳;邵璇;曹海岩;王宁;张珊

参与机构

中国海洋大学;商丘师范学院;山东大学

项目标书摘要:二甲基巯基丙酸内盐(DMSP)年产量高达10亿吨,是海洋中重要的有机硫。海洋细菌对DMSP的代谢是全球碳、硫循环的重要过程,主要包括去甲基化代谢和裂解代谢,其中去甲基化代谢约占DMSP总代谢通量的90%,是DMSP代谢的主要途径。DMSP去甲基化代谢包括四个关键酶:DmdA、DmdB、DmdC和DmdD,其中DmdB、DmdC的结构及催化机制至今未被揭示。本项目以代谢DMSP主要细菌类群——玫瑰杆菌类群细菌为材料,开展如下研究:(1)解析DmdB及其与底物、产物复合物的结构,结合生化研究,揭示其催化MMPA与CoA反应生成MMPA-CoA的分子机制。(2)解析DmdC及其与底物、产物复合物的结构,结合生化研究,揭示其催化MMPA-CoA脱氢生成MTA-CoA的分子机制。上述研究结果将完善海洋细菌参与的DMSP去甲基化代谢的分子机制,有助于更好的认识地球碳、硫循过程,具有重要的理论意义。

Application Abstract: Dimethylsulfoniopropionate(DMSP),produced on a scale of 1000 Tg a year,is an important organic sulfur in the ocean.Microbial cleavage of DMSP is an important step in the global sulfur and carbon cycles.Most DMSP is degraded by marine bacteria via either a demethylation pathway or a cleavage pathway.Between them,demethylation pathway which accounts for about 90%of total metabolism of DMSP,is the major metabolism pathway.There are four key enzymes in the demethylation pathway,DmdA,DmdB,DmdC,and DmdD.However,till now,the structures and catalytic mechanisms of DmdB and DmdC have not been revealed.Roseobacter is one of the major bacterial groups that metabolize DMSP.This program taking Roseobacter as the material will carry out the following studies:(1)Solve the structures of DmdB,DmdB in complex with substrate/product.By combining structural analysis with biochemical experiments,reveal the molecular mechanism that DmdB catalyzes the reaction of MMPA and CoA to produce MMPA-CoA.(2)Solve the structures of DmdC,DmdC in complex with substrate/product.By combining structural analysis with biochemical experiments,reveal the molecular mechanism that DmdC catalyzes the dehydrogenation of MMPA-CoA to MTA-CoA.The research results will consummate the molecular mechanism of DMSP demethylation metabolism and help us to better understand the process of carbon and sulfur cyclings on the earth.

项目受资助省

山东省

项目结题报告(全文)

二甲基巯基丙酸内盐(Dimethylsulfoniopropionate,DMSP)年产量高达10亿吨,是全球硫循环的重要载体物质。微生物介导的DMSP的分解代谢是全球硫循环和碳循环的重要步骤,主要包括去甲基化代谢和裂解代谢。其中去甲基化代谢约占DMSP总代谢通量的90%,是DMSP代谢的主要途径。DMSP去甲基化代谢包括四个关键酶:DmdA、DmdB、DmdC和DmdD,其中DmdB和DmdC的结构及催化机制至今未被揭示。本项目:(1)购买、培养并保藏了典型的DMSP代谢细菌,为DMSP代谢相关研究提供了菌株保障。(2)从体内、体外两个层面验证了典型DMSP代谢细菌中DmdB和DmdC的功能。(3)异源表达纯化了DmdB和DmdC并检测了他们的酶学性质。(4)对DmdB和DmdC并进行了晶体筛选,解析了DmdB和DmdC的晶体结构,并在结构分析和突变验证的基础上阐明了DmdB催化MMPA与CoA反应生成MMPA-CoA的分子机制和DmdC催化MMPA-CoA脱氢生成MTA-CoA的分子机制。(5)对DMSP去甲基化代谢途径中各个酶与其底物的亲和力进行了分析,提出了DMSP去甲基化代谢的动力学调控机制。项目的实施为更好的认识微生物参与的DMSP的去甲基化代谢过程提供了重要帮助。在本项目资助下共发表SCI论文12篇,其中在海洋科学、微生物学和生物化学主流杂志上发表SCI论文10篇。

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  • 1.Genome sequencing and comparative genomics analysis of Halomonas sp. MT13 reveal genetic adaptation to deep-sea environment

    • 关键词:
    • Genome; Comparative genomics; Halomonas sp. MT13; Deep-sea environmentaladaptation;HYDROTHERMAL-VENT; SP-NOV.; BACTERIA; GENUS
    • Wan, Jin-Jian;Wang, Fan;Zhang, Xi-Ying;Xin, Yu;Tian, Ji-Wei;Zhang, Yu-Zhong;Li, Chun-Yang;Fu, Hui-Hui
    • 《MARINE GENOMICS》
    • 2022年
    • 61卷
    • 期刊

    Halomonas sp. MT13, a moderately psychrotolerant, piezotolerant and exopolysaccharide-producing bacterium, was isolated from deep-sea sediment of the Mariana Trench at the depth of 8300 m. Here, we report the complete genome sequence of strain MT13 and its genomic characteristics related to deep-sea environmental adaptation by comparing with its three closely related Halomonas species. The genome of strain MT13 contains one circular chromosome of 3,643,760 bp without any plasmid. Gene annotation, Cluster of Orthologous Groups (COG) and KEGG analysis showed that strain MT13 possesses a serial of genes involved in deep-sea environmental adaptation, including ectoine biosynthesis, osmolyte transport, and cold-shock response. Compared with type strains of three closely related Halomonas species, strain MT13 has higher proportions of genes assigned to translation, ribosomal structure and biogenesis, and coenzyme, lipid and inorganic ion transport and metabolism, but lacks genes involved in flagellar assembly. The genome of strain MT13 would deepen our knowledge on the adaptation strategies of microorganisms dwelling in deep-sea environment.

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  • 2.D-Alanine Metabolism via D-Ala Aminotransferase by a Marine Gammaproteobacterium, Pseudoalteromonas sp. Strain CF6-2

    • 关键词:
    • D-alanine; D-Ala; D-alanine catabolism; D-alanine aminotransferase;alanine racemase; marine bacteria;DISSOLVED ORGANIC-MATTER; D-AMINO-ACIDS; NITROGEN; RACEMASE; ISOMERS;ORIGIN; CARBON; OCEAN; OLD
    • Yu, Yang;Yang, Jie;Teng, Zhao-Jie;Zheng, Li-Yuan;Sheng, Qi;Li, Ping-Yi;Fu, Hui-Hui;Li, Chun-Yang;Chen, Yin;Zhang, Yu-Zhong;Ding, Jun-Mei;Chen, Xiu-Lan
    • 《APPLIED AND ENVIRONMENTAL MICROBIOLOGY》
    • 2022年
    • 88卷
    • 3期
    • 期刊

    As the most abundant D-amino acid (DAA) in the ocean, D-alanine (D-Ala) is a key component of peptidoglycan in the bacterial cell wall. However, the underlying mechanisms of bacterial metabolization of D-Ala through the microbial food web remain largely unknown. In this study, the metabolism of D-Ala by marine bacterium Pseudoalteromonas sp. strain CF6-2 was investigated. Based on genomic, transcriptional, and biochemical analyses combined with gene knockout, D-Ala aminotransferase was found to be indispensable for the catabolism of D-Ala in strain CF6-2. Investigation on other marine bacteria also showed that D-Ala aminotransferase gene is a reliable indicator for their ability to utilize D-Ala. Bioinformatic investigation revealed that D-Ala aminotransferase sequences are prevalent in genomes of marine bacteria and metagenomes, especially in seawater samples, and Gammaproteobacteria represents the predominant group containing D-Ala aminotransferase. Thus, Gammaproteobacteria is likely the dominant group to utilize D-Ala via D-Ala aminotransferase to drive the recycling and mineralization of D-Ala in the ocean.IMPORTANCE As the most abundant D-amino acid in the ocean, D-Ala is a component of the marine DON (dissolved organic nitrogen) pool. However, the underlying mechanism of bacterial metabolization of D-Ala to drive the recycling and mineralization of D-Ala in the ocean is still largely unknown. The results in this study showed that D-Ala aminotransferase is specific and indispensable for D-Ala catabolism in marine bacteria and that marine bacteria containing D-Ala aminotransferase genes are predominantly Gammaproteobacteria widely distributed in global oceans. This study reveals marine D-Ala-utilizing bacteria and the mechanism of their metabolization of D-Ala. The results shed light on the mechanisms of recycling and mineralization of D-Ala driven by bacteria in the ocean, which are helpful in understanding oceanic microbial-mediated nitrogen cycle.

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  • 3.Halomonas profundi sp. nov., isolated from deep- sea sediment of the Mariana Trench

    • 关键词:
    • Halomonas profundi sp; nov; polyphasic taxonomy; deep; sea sediment;Mariana Trench;SALT-LAKE; BACTERIUM; SEQUENCES; STRAINS; FAMILY; GENUS
    • Wang, Fan;Wan, Jin-Jian;Zhang, Xi-Ying;Xin, Yu;Sun, Mei-Ling;Wang, Peng;Zhang, Wei-Peng;Tian, Ji-Wei;Zhang, Yu-Zhong;Li, Chun-Yang;Fu, Hui-Hui
    • 《INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY》
    • 2022年
    • 72卷
    • 1期
    • 期刊

    Two novel Gram- stain- negative, facultative anaerobic, non- flagellated, rod- shaped bacterial strains, designated MT13T and MT32, were isolated from sediment samples collected from the Mariana Trench at a depth of 8300 m. The two strains grew at -2-30 degrees C (optimum, 25 degrees C), at pH 5.5-10.0 (optimum, pH 7.5-8.0) and with 0-15 % (w/v) NaCl (optimum, 3-6 %). They did not reduce nitrate to nitrite nor hydrolyse Tweens 40 and 80, aesculin, casein, starch and DNA. The genomic G+C contents of draft genomes of strain MT13T and MT32 were 52.2 and 54.1 m ol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strains MT13T and MT32 were affiliated with the genus Halomonas, with the highest similarity to the type strain of Halomonas olivaria. The values of average nucleotide identity and in silico DNA-DNA hybridization between strain MT13T and MT32, and between strain MT13T and five closely related type strains of Halomonas species indicated that strains MT13T and MT32 belonged to the same species, but represented a novel species in the genus of Halomonas. The major cellular fatty acids of strains MT13T and MT32 were C16:0, summed feature 3(C16: 1 omega 7c/omega 6c) and summed feature 8 (C18:1 omega 7c/omega 6c). Major polar lipids of strains MT13T and MT32 included phosphatidylglycerol, phosphatidylethanolamine and diphosphatidylglycerol. Ubiquinone- 9 was the predominant respiratory quinone. Based on data from the present polyphasic study, strains MT13T and MT32 represent a novel species of the genus Halomonas, for which the name Halomonas profundi sp. nov. is proposed. The type strain is MT13T (=MCCC 1K06389T=KCTC 82923T).

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  • 4.Novel Insights into Dimethylsulfoniopropionate Catabolism by Cultivable Bacteria in the Arctic Kongsfjorden

    • 关键词:
    • the Arctic; DMSP; marine bacteria; catabolic pathways;Gammaproteobacteria;DIMETHYL SULFIDE; DIMETHYLSULPHONIOPROPIONATE; SULFUR; DMSP; MOLECULE;GAS
    • Zhang, Shan;Cao, Hai-Yan;Zhang, Nan;Teng, Zhao-Jie;Yu, Yang;Wang, Zhi-Bin;Wang, Peng;Fu, Hui-Hui;Chen, Xiu-Lan;Zhang, Yu-Zhong;Li, Chun-Yang
    • 《APPLIED AND ENVIRONMENTAL MICROBIOLOGY》
    • 2022年
    • 88卷
    • 2期
    • 期刊

    Dimethylsulfoniopropionate (DMSP) is one of the most abundant organic sulfur compounds in the oceans, which is mainly degraded by bacteria through two pathways, a cleavage pathway and a demethylation pathway. Its volatile catabolites dimethyl sulfide (DMS) and methanethiol (MT) in these pathways play important roles in the global sulfur cycle and have potential influences on the global climate. Intense DMS/DMSP cycling occurs in the Arctic. However, little is known about the diversity of cultivable DMSP-catabolizing bacteria in the Arctic and how they catabolize DMSP. Here, we screened DMSP-catabolizing bacteria from Arctic samples and found that bacteria of four genera (Psychrobacter, Pseudoalteromonas, Alteromonas, and Vibrio) could grow with DMSP as the sole carbon source, among which Psychrobacter and Pseudoalteromonas are predominant. Four representative strains (Psychrobacter sp. K31L, Pseudoalteromonas sp. K222D, Alteromonas sp. K632G, and Vibrio sp. G41H) from different genera were selected to probe their DMSP catabolic pathways. All these strains produce DMS and MT simultaneously during their growth on DMSP, indicating that all strains likely possess the two DMSP catabolic pathways. On the basis of genomic and biochemical analyses, the DMSP catabolic pathways in these strains were proposed. Bioinformatic analysis indicated that most Psychrobacter and Vibrio bacteria have the potential to catabolize DMSP via the demethylation pathway and that only a small portion of Psychrobacter strains may catabolize DMSP via the cleavage pathway. This study provides novel insights into DMSP catabolism in marine bacteria.IMPORTANCE Dimethylsulfoniopropionate (DMSP) is abundant in the oceans. The catabolism of DMSP is an important step of the global sulfur cycle. Although Gammaproteobacteria are widespread in the oceans, the contribution of Gammaproteobacteria in global DMSP catabolism is not fully understood. Here, we found that bacteria of four genera belonging to Gammaproteobacteria (Psychrobacter, Pseudoalteromonas, Alteromonas and Vibrio), which were isolated from Arctic samples, were able to grow on DMSP. The DMSP catabolic pathways of representative strains were proposed. Bioinformatic analysis indicates that most Psychrobacter and Vibrio bacteria have the potential to catabolize DMSP via the demethylation pathway and that only a small portion of Psychrobacter strains may catabolize DMSP via the cleavage pathway. Our results suggest that novel DMSP dethiomethylases/demethylases may exist in Pseudoalteromonas, Alteromonas, and Vibrio and that Gammaproteobacteria may be important participants in the marine environment, especially in polar DMSP cycling.

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  • 6.Tritonibacter aquimaris sp. nov. and Tritonibacter litoralis sp. nov., two novel members of the Roseobacter group isolated from coastal seawater

    • 关键词:
    • Tritonibacter aquimaris sp; nov; Tritonibacter litoralis sp; nov; TheRoseobacter group; Polyphasic taxonomy; Genomic sequencing and analysis;DMSP lyases;MULTIPLE SEQUENCE ALIGNMENT; DIMETHYLSULFONIOPROPIONATE; CATABOLISM;MICROORGANISMS; COMBINATIONS; BACTERIA; DIVERSE; TREES; NAMES
    • Li, Na;He, Xiao-yan;Liu, Ning-hua;Gu, Tie-ji;Li, Jian;Geng, Yu-hui;Zhang, Shan;Wang, Peng;Fu, Hui-hui;Shi, Mei;Chen, Xiu-lan;Zhang, Yu-zhong;Zhang, Xi-ying;Qin, Qi-long
    • 《ANTONIE VAN LEEUWENHOEK INTERNATIONAL JOURNAL OF GENERAL AND MOLECULARMICROBIOLOGY》
    • 2021年
    • 114卷
    • 6期
    • 期刊

    Two Gram-stain-negative bacterial strains, SM1969(T) and SM1979(T), were isolated from coastal surface seawater of Qingdao, China. They were taxonomically characterized by the phylogenetic, genomic, chemotaxonomic and phenotypic analyses. The two strains shared 97.0% 16S rRNA gene sequence similarity with each other and the highest similarity (96.8-97.5%) with type strains of six species in the genera Shimia, Tritonibacter and Tropicibacter in the Roseobacter group of the family Rhodobacteraceae. In the phylogenetic tree based on single-copy orthologous clusters (OCs), both strains clustered with known species of the genus Tritonibacter and together formed a separate branch adjacent to Tritonibacter ulvae. Although sharing many chemotaxonomic and phenotypic characteristics, the two strains could be differentiated from each other and closely related species by numerous traits. Particularly, strain SM1969(T) was found to have a DMSP lyase coding gene dddW in its genome and have the ability to produce DMS from DMSP while strain SM1979(T) was not. The average nucleotide identity and in silico DNA-DNA hybridization values between strains SM1969(T) and SM1979(T) and type strains of closely related species were all below the thresholds to discriminate bacterial species, demonstrating that they constitute two new species in the genus Tritonibacter. The names Tritonibacter aquimaris sp. nov. and Tritonibacter litoralis sp. nov. are proposed for the two new species, with type strains being SM1969(T) (= MCCC 1K04320(T) = KCTC 72843(T)) and SM1979(T) (= MCCC 1K04321(T) = KCTC 72842(T)), respectively.

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  • 7.Comparison of Alginate Utilization Pathways in Culturable Bacteria Isolated From Arctic and Antarctic Marine Environments

    • 关键词:
    • alginate; polar bacteria; enzymes; transporters; biogeographicdistribution;HUMAN GUT SYMBIONT; SP NOV.; BACTEROIDES-THETAIOTAOMICRON;MOLECULAR-IDENTIFICATION; GEN. NOV.; STARCH; LYASE; CATABOLISM;ALIGNMENT; PROTEIN
    • Cha, Qian-Qian;Wang, Xiu-Juan;Ren, Xue-Bing;Li, Dong;Wang, Peng;Li, Ping-Yi;Fu, Hui-Hui;Zhang, Xi-Ying;Chen, Xiu-Lan;Zhang, Yu-Zhong;Xu, Fei;Qin, Qi-Long
    • 《FRONTIERS IN MICROBIOLOGY》
    • 2021年
    • 12卷
    • 期刊

    Alginate, mainly derived from brown algae, is an important carbon source that can support the growth of marine microorganisms in the Arctic and Antarctic regions. However, there is a lack of systematic investigation and comparison of alginate utilization pathways in culturable bacteria from both polar regions. In this study, 88 strains were isolated from the Arctic and Antarctic regions, of which 60 strains could grow in the medium with alginate as the sole carbon source. These alginate-utilizing strains belong to 9 genera of the phyla Proteobacteria and Bacteroidetes. The genomes of 26 alginate-utilizing strains were sequenced and genomic analyses showed that they all contain the gene clusters related to alginate utilization. The alginate transport systems of Proteobacteria differ from those of Bacteroidetes and there may be unique transport systems among different genera of Proteobacteria. The biogeographic distribution pattern of alginate utilization genes was further investigated. The alginate utilization genes are found to cluster according to bacterial taxonomy rather than geographic location, indicating that the alginate utilization genes do not evolve independently in both polar regions. This study systematically illustrates the alginate utilization pathways in culturable bacteria from the Arctic and Antarctic regions, shedding light into the distribution and evolution of alginate utilization pathways in polar bacteria.

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  • 8.Biogeographic traits of dimethyl sulfide and dimethylsulfoniopropionate cycling in polar oceans

    • 关键词:
    • Polar oceans; DMS; DMSP cycling; Geographic distribution; Phylogeneticdiversity;PARTICULATE DIMETHYLSULFOXIDE; DEPENDENT DEMETHYLASE; MISSING LINK;DMSP; DIMETHYLSULPHONIOPROPIONATE; LYASE; SEA; REDUCTASE; MECHANISM;COASTAL
    • Teng, Zhao-Jie;Qin, Qi-Long;Zhang, Weipeng;Li, Jian;Fu, Hui-Hui;Wang, Peng;Lan, Musheng;Lu, Guangfu;He, Jianfeng;McMinn, Andrew;Wang, Min;Chen, Xiu-Lan;Zhang, Yu-Zhong;Chen, Yin;Li, Chun-Yang
    • 《MICROBIOME》
    • 2021年
    • 9卷
    • 1期
    • 期刊

    Background: Dimethyl sulfide (DMS) is the dominant volatile organic sulfur in global oceans. The predominant source of oceanic DMS is the cleavage of dimethylsulfoniopropionate (DMSP), which can be produced by marine bacteria and phytoplankton. Polar oceans, which represent about one fifth of Earth's surface, contribute significantly to the global oceanic DMS sea-air flux. However, a global overview of DMS and DMSP cycling in polar oceans is still lacking and the key genes and the microbial assemblages involved in DMSP/DMS transformation remain to be fully unveiled.Results: Here, we systematically investigated the biogeographic traits of 16 key microbial enzymes involved in DMS/DMSP cycling in 60 metagenomic samples from polar waters, together with 174 metagenome and 151 metatranscriptomes from non-polar Tara Ocean dataset. Our analyses suggest that intense DMS/DMSP cycling occurs in the polar oceans. DMSP demethylase (DmdA), DMSP lyases (DddD, DddP, and DddK), and trimethylamine monooxygenase (Tmm, which oxidizes DMS to dimethylsulfoxide) were the most prevalent bacterial genes involved in global DMS/DMSP cycling. Alphaproteobacteria (Pelagibacterales) and Gammaproteobacteria appear to play prominent roles in DMS/DMSP cycling in polar oceans. The phenomenon that multiple DMS/DMSP cycling genes co-occurred in the same bacterial genome was also observed in metagenome assembled genomes (MAGs) from polar oceans. The microbial assemblages from the polar oceans were significantly correlated with water depth rather than geographic distance, suggesting the differences of habitats between surface and deep waters rather than dispersal limitation are the key factors shaping microbial assemblages involved in DMS/DMSP cycling in polar oceans.Conclusions: Overall, this study provides a global overview of the biogeographic traits of known bacterial genes involved in DMS/DMSP cycling from the Arctic and Antarctic oceans, laying a solid foundation for further studies of DMS/DMSP cycling in polar ocean microbiome at the enzymatic, metabolic, and processual levels.

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  • 9.A novel ATP dependent dimethylsulfoniopropionate lyase in bacteria that releases dimethyl sulfide and acryloyl-CoA

    • 关键词:
    • MARINE-BACTERIA; ACETYL-COENZYME; MOLECULAR INSIGHT; DMSP; SULFUR;SYNTHETASES; GENE; DIMETHYLSULPHONIOPROPIONATE; IDENTIFICATION;BIOSYNTHESIS
    • Li, Chun-Yang;Wang, Xiu-Juan;Chen, Xiu-Lan;Sheng, Qi;Zhang, Shan;Wang, Peng;Quareshy, Mussa;Rihtman, Branko;Shao, Xuan;Gao, Chao;Li, Fuchuan;Li, Shengying;Zhang, Weipeng;Zhang, Xiao-Hua;Yang, Gui-Peng;Todd, Jonathan D.;Chen, Yin;Zhang, Yu-Zhong
    • 《ELIFE》
    • 2021年
    • 10卷
    • 期刊

    Dimethylsulfoniopropionate (DMSP) is an abundant and ubiquitous organosulfur molecule in marine environments with important roles in global sulfur and nutrient cycling. Diverse DMSP lyases in some algae, bacteria, and fungi cleave DMSP to yield gaseous dimethyl sulfide (DMS), an infochemical with important roles in atmospheric chemistry. Here, we identified a novel ATP-dependent DMSP lyase, DddX. DddX belongs to the acyl-CoA synthetase superfamily and is distinct from the eight other known DMSP lyases. DddX catalyses the conversion of DMSP to DMS via a two--step reaction: the ligation of DMSP with CoA to form the intermediate DMSP-CoA, which is then cleaved to DMS and acryloyl-CoA. The novel catalytic mechanism was elucidated by structural and biochemical analyses. DddX is found in several Alphaproteobacteria, Gammaproteobacteria, and Firmicutes, suggesting that this new DMSP lyase may play an overlooked role in DMSP/DMS cycles.

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