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321.
322.
Sebastian Peter Matthias Kinne René Ullrich Gernot Kayser Martin Hofrichter 《Enzyme and microbial technology》2013,52(6-7):370-376
Unspecific peroxygenases (EC 1.11.2.1) represent a group of secreted heme-thiolate proteins that are capable of catalyzing the mono-oxygenation of diverse organic compounds, using only H2O2 as a co-substrate. Here we show that the peroxygenase secreted by the fungus Agrocybe aegerita catalyzed the oxidation of 20 different alkenes. Five branched alkenes, among them 2,3-dimethyl-2-butene and cis-2-butene, as well as propene and butadiene were epoxidized with complete regioselectivity. Longer linear alkenes with a terminal double bond (e.g. 1-octene) and cyclic alkenes (e.g. cyclohexene) were converted into the corresponding epoxides and allylic hydroxylation products; oxidation of the cyclic monoterpene limonene yielded three oxygenation products (two epoxides and an alcohol). In the case of 1-alkenes, the conversion occurred with moderate stereoselectivity, in which the preponderance for the (S)-enantiomer reached up to 72% ee for the epoxide product. The apparent Michaelis–Menten constant (Km) for the epoxidation of the model substrate 2-methyl-2-butene was 5 mM, the turnover number (kcat) 1.3 × 103 s?1 and the calculated catalytic efficiency, kcat/Km, was 2.5 × 105 M?1 s?1. As epoxides represent chemical building blocks of high relevance, new enzymatic epoxidation pathways are of interest to complement existing chemical and biotechnological approaches. Stable and versatile peroxygenases as that of A. aegerita may form a promising biocatalytic platform for the development of such enzyme-based syntheses. 相似文献
323.
Honglei Jia Jisheng Li Jingen Zhu Tingting Fan Dong Qian Yuelong Zhou Jiaojiao Wang Haiyun Ren Yun Xiang Lizhe An 《The Journal of biological chemistry》2013,288(45):32277-32288
Higher order actin filament structures are necessary for cytoplasmic streaming, organelle movement, and other physiological processes. However, the mechanism by which the higher order cytoskeleton is formed in plants remains unknown. In this study, we identified a novel actin-cross-linking protein family (named CROLIN) that is well conserved only in the plant kingdom. There are six isovariants of CROLIN in the Arabidopsis genome, with CROLIN1 specifically expressed in pollen. In vitro biochemical analyses showed that CROLIN1 is a novel actin-cross-linking protein with binding and stabilizing activities. Remarkably, CROLIN1 can cross-link actin bundles into actin networks. CROLIN1 loss of function induces pollen germination and pollen tube growth hypersensitive to latrunculin B. All of these results demonstrate that CROLIN1 may play an important role in stabilizing and remodeling actin filaments by binding to and cross-linking actin filaments. 相似文献
324.
325.
Huan Yan Bo Peng Wenhui He Guocai Zhong Yonghe Qi Bijie Ren Zhenchao Gao Zhiyi Jing Mei Song Guangwei Xu Jianhua Sui Wenhui Li 《Journal of virology》2013,87(14):7977-7991
Human hepatitis B virus (HBV) and its satellite virus, hepatitis D virus (HDV), primarily infect humans, chimpanzees, or tree shrews (Tupaia belangeri). Viral infections in other species are known to be mainly restricted at the entry level since viral replication can be achieved in the cells by transfection of the viral genome. Sodium taurocholate cotransporting polypeptide (NTCP) is a functional receptor for HBV and HDV, and amino acids 157 to 165 of NTCP are critical for viral entry and likely limit viral infection of macaques. However, the molecular determinants for viral entry restriction in mouse NTCP (mNTCP) remain unclear. In this study, mNTCP was found to be unable to support either HBV or HDV infection, although it can bind to pre-S1 of HBV L protein and is functional in transporting substrate taurocholate; comprehensive swapping and point mutations of human NTCP (hNTCP) and mNTCP revealed molecular determinants restricting mNTCP for viral entry of HBV and HDV. Remarkably, when mNTCP residues 84 to 87 were substituted by human counterparts, mNTCP can effectively support viral infections. In addition, a number of cell lines, regardless of their species or tissue origin, supported HDV infection when transfected with hNTCP or mNTCP with residues 84 to 87 replaced by human counterparts, highlighting the central role of NTCP for viral infections mediated by HBV envelope proteins. These studies advance our understanding of NTCP-mediated viral entry of HBV and HDV and have important implications for developing the mouse model for their infections. 相似文献
326.
Nicholas C. Wu Arthur P. Young Sugandha Dandekar Hemani Wijersuriya Laith Q. Al-Mawsawi Ting-Ting Wu Ren Sun 《Journal of virology》2013,87(2):1193-1199
Compensatory mutations contribute to the appearance of the oseltamivir resistance substitution H274Y in the neuraminidase (NA) gene of H1N1 influenza viruses. Here, we describe a high-throughput screening method utilizing error-prone PCR and next-generation sequencing to comprehensively screen NA genes for H274Y compensatory mutations. We found four mutations that can either fully (R194G, E214D) or partially (L250P, F239Y) compensate for the fitness deficiency of the H274Y mutant. The compensatory effect of E214D is applicable in both seasonal influenza virus strain A/New Caledonia/20/1999 and 2009 pandemic swine influenza virus strain A/California/04/2009. The technique described here has the potential to profile a gene at the single-nucleotide level to comprehend the dynamics of mutation space and fitness and thus offers prediction power for emerging mutant species. 相似文献
327.
Ken'ichiro Matsumoto Yoshikazu Tanaka Tsuyoshi Watanabe Ren Motohashi Koji Ikeda Kota Tobitani Min Yao Isao Tanaka Seiichi Taguchi 《Applied and environmental microbiology》2013,79(19):6134-6139
NADPH-dependent acetoacetyl-coenzyme A (acetoacetyl-CoA) reductase (PhaB) is a key enzyme in the synthesis of poly(3-hydroxybutyrate) [P(3HB)], along with β-ketothiolase (PhaA) and polyhydroxyalkanoate synthase (PhaC). In this study, PhaB from Ralstonia eutropha was engineered by means of directed evolution consisting of an error-prone PCR-mediated mutagenesis and a P(3HB) accumulation-based in vivo screening system using Escherichia coli. From approximately 20,000 mutants, we obtained two mutant candidates bearing Gln47Leu (Q47L) and Thr173Ser (T173S) substitutions. The mutants exhibited kcat values that were 2.4-fold and 3.5-fold higher than that of the wild-type enzyme, respectively. In fact, the PhaB mutants did exhibit enhanced activity and P(3HB) accumulation when expressed in recombinant Corynebacterium glutamicum. Comparative three-dimensional structural analysis of wild-type PhaB and highly active PhaB mutants revealed that the beneficial mutations affected the flexibility around the active site, which in turn played an important role in substrate recognition. Furthermore, both the kinetic analysis and crystal structure data supported the conclusion that PhaB forms a ternary complex with NADPH and acetoacetyl-CoA. These results suggest that the mutations affected the interaction with substrates, resulting in the acquirement of enhanced activity. 相似文献
328.
E.-N. Yang G. M. Rosewarne S. A. Herrera-Foessel J. Huerta-Espino Z.-X. Tang C.-F. Sun Z.-L. Ren R. P. Singh 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2013,126(7):1721-1732
Chapio is a spring wheat developed by CIMMYT in Mexico by a breeding program that focused on multigenic resistances to leaf rust and stripe rust. A population consisting of 277 recombinant inbred lines (RILs) was developed by crossing Chapio with Avocet. The RILs were genotyped with DArT markers (137 randomly selected RILs) and bulked segregant analysis conducted to supplement the map with informative SSR markers. The final map consisted of 264 markers. Phenotyping against stripe rust was conducted for three seasons in Toluca, Mexico and at three sites over two seasons (total of four environments) in Sichuan Province, China. Significant loci across the two inter-continental regions included Lr34/Yr18 on 7DS, Sr2/Yr30 on 3BS, and a QTL on 3D. There were significant genotype × environment interactions with resistance gene Yr31 on 2BS being effective in most of the Toluca environments; however, a late incursion of a virulent pathotype in 2009 rendered this gene ineffective. This locus also had no effect in China. Conversely, a 5BL locus was only effective in the Chinese environments. There were also complex additive interactions. In the Mexican environments, Yr31 suppressed the additive effect of Yr30 and the 3D locus, but not of Lr34/Yr18, while in China, the 3D and 5BL loci were generally not additive with each other, but were additive when combined with other loci. These results indicate the importance of maintaining diverse, multi-genic resistances as Chapio had stable inter-continental resistance despite the fact that there were QTLs that were not effective in either one or the other region. 相似文献
329.
330.
Jiachuan Pan Xin Xie Wang Tian Ali Adem Bahar Nan Lin Fangchao Song Jing An Dacheng Ren 《Applied microbiology and biotechnology》2013,97(20):9145-9154
Persisters are a small subpopulation of bacterial cells that are dormant and extremely tolerant to antibiotics. The intrinsic antibiotic tolerance of persisters also facilitates the development of multidrug resistance through acquired mechanisms based on drug resistance genes. In this study, we demonstrate that (Z)-4-bromo-5-(bromomethylene)-3-methylfuran-2(5H)-one (BF8) can reduce persistence during Escherichia coli growth and revert the antibiotic tolerance of its persister cells. The effects of BF8 were more profound when the pH was increased from 6 to 8.5. Although BF8 is a quorum sensing (QS) inhibitor, similar effects were observed for the wild-type E. coli RP437 and its ΔluxS mutant, suggesting that these effects did not occur solely through inhibition of AI-2-mediated QS. In addition to its effects on planktonic persisters, BF8 was also found to disperse RP437 biofilms and to render associated cells more sensitive to ofloxacin. At the doses that are effective against E. coli persister cells, BF8 appeared to be safe to the tested normal mammalian cells in vitro and exhibited no long-term cytotoxicity to normal mouse tissues in vivo. These findings broadened the activities of brominated furanones and shed new light on persister control. 相似文献