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21.
Hobbs JK Shepherd C Saul DJ Demetras NJ Haaning S Monk CR Daniel RM Arcus VL 《Molecular biology and evolution》2012,29(2):825-835
Thermophily is thought to be a primitive trait, characteristic of early forms of life on Earth, that has been gradually lost over evolutionary time. The genus Bacillus provides an ideal model for studying the evolution of thermophily as it is an ancient taxon and its contemporary species inhabit a range of thermal environments. The thermostability of reconstructed ancestral proteins has been used as a proxy for ancient thermal adaptation. The reconstruction of ancestral "enzymes" has the added advantages of demonstrable activity, which acts as an internal control for accurate inference, and providing insights into the evolution of enzymatic catalysis. Here, we report the reconstruction of the structurally complex core metabolic enzyme LeuB (3-isopropylmalate dehydrogenase, E. C. 1.1.1.85) from the last common ancestor (LCA) of Bacillus using both maximum likelihood (ML) and Bayesian inference. ML LeuB from the LCA of Bacillus shares only 76% sequence identity with its closest contemporary homolog, yet it is fully functional, thermophilic, and exhibits high values for k(cat), k(cat)/K(M), and ΔG(?) for unfolding. The Bayesian version of this enzyme is also thermophilic but exhibits anomalous catalytic kinetics. We have determined the 3D structure of the ML enzyme and found that it is more closely aligned with LeuB from deeply branching bacteria, such as Thermotoga maritima, than contemporary Bacillus species. To investigate the evolution of thermophily, three descendents of LeuB from the LCA of Bacillus were also reconstructed. They reveal a fluctuating trend in thermal evolution, with a temporal adaptation toward mesophily followed by a more recent return to thermophily. Structural analysis suggests that the determinants of thermophily in LeuB from the LCA of Bacillus and the most recent ancestor are distinct and that thermophily has arisen in this genus at least twice via independent evolutionary paths. Our results add significant fluctuations to the broad trend in thermal adaptation previously proposed and demonstrate that thermophily is not exclusively a primitive trait, as it can be readily gained as well as lost. Our findings also demonstrate that reconstruction of complex functional Precambrian enzymes is possible and can provide empirical access to the evolution of ancient phenotypes and metabolisms. 相似文献
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23.
Mogens K. Boisen Christian Dehlendorff Dorte Linnemann Boye S. Nielsen Jim S. Larsen Kell ?sterlind Svend E. Nielsen Line S. Tarpgaard Camilla Qvortrup Per Pfeiffer Niels H. Holl?nder Nina Keldsen Torben F. Hansen Brita B. Jensen Estrid V. S. H?gdall Benny V. Jensen Julia S. Johansen 《PloS one》2014,9(10)
Purpose
We tested the hypothesis that expression of microRNAs (miRNAs) in cancer tissue can predict effectiveness of bevacizumab added to capecitabine and oxaliplatin (CAPEOX) in patients with metastatic colorectal cancer (mCRC).Experimental Design
Patients with mCRC treated with first line CAPEOX and bevacizumab (CAPEOXBEV): screening (n = 212) and validation (n = 121) cohorts, or CAPEOX alone: control cohort (n = 127), were identified retrospectively and archival primary tumor samples were collected. Expression of 754 miRNAs was analyzed in the screening cohort using polymerase chain reaction (PCR) arrays and expression levels were related to time to disease progression (TTP) and overall survival (OS). Significant miRNAs from the screening study were analyzed in all three cohorts using custom PCR arrays. In situ hybridization (ISH) was done for selected miRNAs.Results
In the screening study, 26 miRNAs were significantly correlated with outcome in multivariate analyses. Twenty-two miRNAs were selected for further study. Higher miR-664-3p expression and lower miR-455-5p expression were predictive of improved outcome in the CAPEOXBEV cohorts and showed a significant interaction with bevacizumab effectiveness. The effects were strongest for OS. Both miRNAs showed high expression in stromal cells. Higher expression of miR-196b-5p and miR-592 predicted improved outcome regardless of bevacizumab treatment, with similar effect estimates in all three cohorts.Conclusions
We have identified potentially predictive miRNAs for bevacizumab effectiveness and additional miRNAs that could be related to chemotherapy effectiveness or prognosis in patients with mCRC. Our findings need further validation in large cohorts, preferably from completed randomized trials. 相似文献24.
Conticello SG Thomas CJ Petersen-Mahrt SK Neuberger MS 《Molecular biology and evolution》2005,22(2):367-377
The AID/APOBEC family (comprising AID, APOBEC1, APOBEC2, and APOBEC3 subgroups) contains members that can deaminate cytidine in RNA and/or DNA and exhibit diverse physiological functions (AID and APOBEC3 deaminating DNA to trigger pathways in adaptive and innate immunity; APOBEC1 mediating apolipoprotein B RNA editing). The founder member APOBEC1, which has been used as a paradigm, is an RNA-editing enzyme with proposed antecedents in yeast. Here, we have undertaken phylogenetic analysis to glean insight into the primary physiological function of the AID/APOBEC family. We find that although the family forms part of a larger superfamily of deaminases distributed throughout the biological world, the AID/APOBEC family itself is restricted to vertebrates with homologs of AID (a DNA deaminase that triggers antibody gene diversification) and of APOBEC2 (unknown function) identifiable in sequence databases from bony fish, birds, amphibians, and mammals. The cloning of an AID homolog from dogfish reveals that AID extends at least as far back as cartilaginous fish. Like mammalian AID, the pufferfish AID homolog can trigger deoxycytidine deamination in DNA but, consistent with its cold-blooded origin, is thermolabile. The fine specificity of its mutator activity and the biased codon usage in pufferfish IgV genes appear broadly similar to that of their mammalian counterparts, consistent with a coevolution of the antibody mutator and its substrate for the optimal targeting of somatic mutation during antibody maturation. By contrast, APOBEC1 and APOBEC3 are later evolutionary arrivals with orthologs not found in pufferfish (although synteny with mammals is maintained in respect of the flanking loci). We conclude that AID and APOBEC2 are likely to be the ancestral members of the AID/APOBEC family (going back to the beginning of vertebrate speciation) with both APOBEC1 and APOBEC3 being mammal-specific derivatives of AID and a complex set of domain shuffling underpinning the expansion and evolution of the primate APOBEC3s. 相似文献
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26.
Calpe S Erdos E Liao G Wang N Rietdijk S Simarro M Scholtz B Mooney J Lee CH Shin MS Rajnavölgyi E Schatzle J Morse HC Terhorst C Lanyi A 《Immunogenetics》2006,58(1):15-25
Human EAT-2 (SH2D1B) and SLAM-associated protein (SAP) (SH2D1A) are single SH2-domain adapters, which bind to specific tyrosine
residues in the cytoplasmic tail of six signaling lymphocytic activation molecule (SLAM) (SLAMF1)-related receptors. Here
we report that, unlike in humans, the mouse and rat Eat2 genes are duplicated with an identical genomic organization. The coding regions of the mouse Eat2a and Eat2b genes share 91% identity at the nucleotide level and 84% at the protein level; similarly, segments of introns are highly
conserved. Whereas expression of mouse Eat2a mRNA was detected in multiple tissues, Eat2b was only detectable in mouse natural killer cells, CD8+ T cells, and ovaries, suggesting a very restricted tissue expression of the latter. Both the EAT-2A and EAT-2B coimmunoprecipitated
with mouse SLAM in transfected cells and augmented tyrosine phosphorylation of the cytoplasmic tail of SLAM. Both EAT-2A and
EAT-2B bind to the Src-like kinases Fyn, Hck, Lyn, Lck, and Fgr, as determined by a yeast two-hybrid assay. However, unlike
SAP, the EAT-2 proteins bind to their kinase domains and not to the SH3 domain of these kinases. Taken together, the data
suggest that both EAT-2A and EAT-2B are adapters that recruit Src kinases to SLAM family receptors using a mechanism that
is distinct from that of SAP.
Electronic supplementary material Supplementary material is available for this article at and accessible for authorised users.
S. Calpe and E. Erdős contributed equally to this work 相似文献
27.
Lung infection with Pseudomonas aeruginosa, leading to chronic lung disease with impaired function, is the major course of morbidity and mortality among cystic fibrosis patients. The bacterium produces two lectins that bind to alpha-D-galactose (PA-IL) and L-fucose (PA-IIL), respectively, and lectin-carbohydrate interactions may be involved in microbial pathogenicity by creating bacterial adherence to epithelial and endothelial cells. An ideal animal model for P. aeruginosa infection has until now not been established, but the mink seems to be the only animal that has been reported to develop spontaneous P. aeruginosa infections in the airways. Since cystic fibrosis also severely may affect pancreatic function, we incubated sections from mink lungs and pancreas with a medium containing Pseudomonas lectins in order to detect in situ binding of the bacterial lectins. In the lungs, both lectins adhered to seromucinous glands located in the submucosa of the larger bronchi. Additionally, PA-IL reacted with the capillaries in the alveolar walls and with the small blood vessels forming the vasa vasorum around the larger vessels, while PA-IIL marked the goblet cells in the bronchial surface epithelium. In the pancreas, both lectins bound to the epithelium in the excretory ducts, and additionally, PA-IL strongly stained the pancreatic capillaries while PA-IIL staining was noticed in the apical part of acinar cells in the exocrine part of the gland while no lectin reaction could be recorded in the endocrine cells. Judging from the results in the present paper the mink should be considered a suitable model to study P. aeruginosa adherence. 相似文献
28.
N‐glycan maturation mutants in Lotus japonicus for basic and applied glycoprotein research 下载免费PDF全文
Carina T. Pedersen Ian Loke Andrea Lorentzen Sara Wolf Manoj Kamble Sebastian K. Kristensen David Munch Simona Radutoiu Edzard Spillner Peter Roepstorff Morten Thaysen‐Andersen Jens Stougaard Svend Dam 《The Plant journal : for cell and molecular biology》2017,91(3):394-407
Studies of protein N‐glycosylation are important for answering fundamental questions on the diverse functions of glycoproteins in plant growth and development. Here we generated and characterised a comprehensive collection of Lotus japonicusLORE1 insertion mutants, each lacking the activity of one of the 12 enzymes required for normal N‐glycan maturation in the glycosylation machinery. The inactivation of the individual genes resulted in altered N‐glycan patterns as documented using mass spectrometry and glycan‐recognising antibodies, indicating successful identification of null mutations in the target glyco‐genes. For example, both mass spectrometry and immunoblotting experiments suggest that proteins derived from the α1,3‐fucosyltransferase (Lj3fuct) mutant completely lacked α1,3‐core fucosylation. Mass spectrometry also suggested that the Lotus japonicus convicilin 2 was one of the main glycoproteins undergoing differential expression/N‐glycosylation in the mutants. Demonstrating the functional importance of glycosylation, reduced growth and seed production phenotypes were observed for the mutant plants lacking functional mannosidase I, N‐acetylglucosaminyltransferase I, and α1,3‐fucosyltransferase, even though the relative protein composition and abundance appeared unaffected. The strength of our N‐glycosylation mutant platform is the broad spectrum of resulting glycoprotein profiles and altered physiological phenotypes that can be produced from single, double, triple and quadruple mutants. This platform will serve as a valuable tool for elucidating the functional role of protein N‐glycosylation in plants. Furthermore, this technology can be used to generate stable plant mutant lines for biopharmaceutical production of glycoproteins displaying relative homogeneous and mammalian‐like N‐glycosylation features. 相似文献
29.
Stephen J. H. Ashcroft Judy Bunce Martin Lowry Svend E. Hansen Carl J. Hedeskov 《The Biochemical journal》1978,174(2):517-526
Rates of incorporation of [4,5-(3)H]leucine into insulin plus proinsulin, designated ;(pro)insulin', and total protein in rat pancreatic islets were measured. Glucose stimulates rates of total protein and (pro)insulin biosynthesis, but (pro)insulin biosynthesis is stimulated preferentially. Mannose and N-acetylglucosamine also stimulate (pro)insulin and total protein biosynthesis; inosine and dihydroxyacetone stimulate (pro)insulin biosynthesis specifically. Fructose does not stimulate (pro)insulin biosynthesis when tested alone, but does so in the presence of low concentrations of glucose, mannose or N-acetylglucosamine. Many glucose analogues do not stimulate (pro)insulin biosynthesis. Mannoheptulose inhibits synthesis of (pro)insulin and total protein stimulated by glucose or mannose but not by dihydroxyacetone, inosine or N-acetylglucosamine; phloretin (9mum) inhibits N-acetylglucosamine-stimulated (pro)insulin biosynthesis preferentially. The data are in agreement with the view that the same glucose-sensor mechanism may control both insulin release and biosynthesis, and ;substrate-site' model is suggested. The threshold for stimulation of biosynthesis of (pro)insulin and total protein is lower than that found for glucose-stimulated insulin release; moreover the biosynthetic response to an elevation of glucose concentration is slower than that found for insulin release. The physiological implication of these findings is discussed. Caffeine and isobutylmethylxanthine, at concentrations known to increase islet 3':5'-cyclic AMP and potentiate glucose-induced insulin release, were without effect on rates of glucose-stimulated (pro)insulin biosynthesis. 相似文献
30.
Alexandra Vanessa Finsen Thor Ueland Ivar Sjaastad Trine Ranheim Mohammed S. Ahmed Christen P. Dahl Erik T. Askevold Svend Aakhus Cathrine Husberg Arnt E. Fiane Martin Lipp Lars Gullestad Geir Christensen P?l Aukrust Arne Yndestad 《PloS one》2014,9(11)