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Flagellin is the subunit of the bacterial filament, the micrometer-long propeller of a bacterial flagellum. The protein is believed to undergo unfolding for transport through the channel of the filament and to refold in a chamber at the end of the channel before being assembled into the growing filament. We report a thermal unfolding simulation study of S. typhimurium flagellin in aqueous solution as an attempt to gain atomic-level insight into the refolding process. Each molecule comprises two filament-core domains {D0, D1} and two hypervariable-region domains {D2, D3}. D2 can be separated into subdomains D2a and D2b. We observed a similar unfolding order of the domains as reported in experimental thermal denaturation. D2a and D3 exhibited high thermal stability and contained persistent three-stranded β-sheets in the denatured state which could serve as folding cores to guide refolding. A recent mutagenesis study on flagellin stability seems to suggest the importance of the folding cores. Using crude size estimates, our data suggests that the chamber might be large enough for either denatured hypervariable-region domains or filament-core domains, but not whole flagellin; this implicates a two-staged refolding process. 相似文献
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Lin Xiao Can Chen Zhendong Li Sumin Zhu Johan Ck Tay Xi Zhang Shijun Zha Jieming Zeng Wee Kiat Tan Xin Liu Wee Joo Chng Shu Wang 《Cytotherapy》2018,20(3):420-435
Vγ9Vδ2 T cells are a minor subset of lymphocytes in the peripheral blood that has been extensively investigated for their tolerability, safety and anticancer efficacy. A hindrance to the broad application of these cells for adoptive cellular immunotherapy has been attaining clinically appropriate numbers of Vγ9Vδ2 T cells. Furthermore, Vγ9Vδ2 T cells exist at low frequencies among cancer patients. We, therefore, sought to conceive an economical method that allows for a quick and robust large-scale expansion of Vγ9Vδ2 T cells. A two-step protocol was developed, in which peripheral blood mononuclear cells (PBMCs) from healthy donors or cancer patients were activated with Zometa and interleukin (IL)-2, followed by co-culturing with gamma-irradiated, CD64-, CD86- and CD137L-expressing K562 artificial antigen-presenting cells (aAPCs) in the presence of the anti-CD3 antibody OKT3. We optimized the co-culture ratio of K562 aAPCs to immune cells, and migrated this method to a G-Rex cell growth platform to derive clinically relevant cell numbers in a Good Manufacturing Practice (GMP)-compliant manner. We further include a depletion step to selectively remove αβ T lymphocytes. The method exhibited high expansion folds and a specific enrichment of Vγ9Vδ2 T cells. Expanded Vγ9Vδ2 T cells displayed an effector memory phenotype with a concomitant down-regulated expression of inhibitory immune checkpoint receptors. Finally, we ascertained the cytotoxic activity of these expanded cells by using nonmodified and chimeric antigen receptor (CAR)–engrafted Vγ9Vδ2 T cells against a panel of solid tumor cells. Overall, we report an efficient approach to generate highly functional Vγ9Vδ2 T cells in massive numbers suitable for clinical application in an allogeneic setting. 相似文献
25.
Marmoset phylogenetics, conservation perspectives, and evolution of the mtDNA control region 总被引:3,自引:1,他引:2
Tagliaro CH; Schneider MP; Schneider H; Sampaio IC; Stanhope MJ 《Molecular biology and evolution》1997,14(6):674-684
Marmosets (genus Callithrix) are a diverse group of platyrrhine primates
with 13-15 purported taxa, many of them considered endangered.
Morphological analyses constitute most of the basis for recognition of
these forms as distinct taxa. The purpose of this study was to provide a
molecular view, based on mitochondrial control region sequences, of the
evolutionary history of the marmosets, concomitant with a molecular
phylogenetic perspective on species diversity within the group. An
additional purpose was to provide the first comparative examination of a
complete New World monkey control region sequence with those of other
mammals. The phylogenetic analyses provide convincing support for a split
between the Atlantic forest and Amazonian marmosets, with the inclusion of
the pygmy marmoset (Cebuella pygmaea) at the base of the Amazonian clade.
The earliest branch of the Atlantic forest group was C. aurita. In the
Amazonian group, the analyses do not support the recognition of C.
humeralifer and the recently described C mauesi as distinct taxa. They do,
however, support a clear distinction between C. argentata and a strongly
supported mixed clade of C. humeralifer and C. mauesi. In the Atlantic
forest group, the phylogenetic tree suggests mixing between C. penicillata,
C. kuhli, and possibly C. jacchus. Most of the sequence features
characteristic of other mammal control regions were also evident in
marmosets, with the exception that conserved sequence blocks (CSBs) 2 and 3
were not clearly identifiable. Tandem repeat units often associated with
heteroplasmy in a variety of other mammals were not evident in the marmoset
sequences.
相似文献
26.
Persistent nuclear ribosomal DNA sequence polymorphism in the Amelanchier agamic complex (Rosaceae) 总被引:5,自引:0,他引:5
Campbell CS; Wojciechowski MF; Baldwin BG; Alice LA; Donoghue MJ 《Molecular biology and evolution》1997,14(1):81-90
Individual plants of several Amelanchier taxa contain many polymorphic
nucleotide sites in the internal transcribed spacers (ITS) of nuclear
ribosomal DNA (nrDNA). This polymorphism is unusual because it is not
recent in origin and thus has resisted homogenization by concerted
evolution. Amelanchier ITS sequence polymorphism is hypothesized to be the
result of gene flow between two major North American clades resolved by
phylogenetic analysis of ITS sequences. Western North American species plus
A. humilis and A. sanguinea of eastern North America form one clade (A),
and the remaining eastern North American Amelanchier make up clade B. Five
eastern North American taxa are polymorphic at many of the nucleotide sites
where clades A and B have diverged and are thought to be of hybrid origin,
with A. humilis or A. sanguinea as one parent and various members of clade
B as the other parent. Morphological evidence suggests that A. humilis is
one of the parents of one of the polymorphic taxa, a microspecies that we
refer to informally as A. "erecta." Sequences of 21 cloned copies of the
ITS1- 5.8S gene-ITS2 region from one A. "erecta" individual are identical
to A. humilis sequence or to the clade B consensus sequence, or they are
apparent recombinants of A. humilis and clade B ITS repeats. Amelanchier
"erecta" and another polymorphic taxon are suspected to be relatively old
because both grow several hundred kilometers beyond the range of one of
their parents. ITS sequence polymorphisms have apparently persisted in
these two taxa perhaps because of polyploidy and/or agamospermy (asexual
seed production), which are prevalent in the genus.
相似文献
27.
Marieke?Pingen Ramin?Sarrami-Forooshani Annemarie?MJ?Wensing Petra?van Ham Agata?Drewniak Charles?AB?Boucher Teunis?BH?GeijtenbeekEmail author Monique?NijhuisEmail author 《Retrovirology》2014,11(1):113
Background
Different patterns of drug resistance are observed in treated and therapy naïve HIV-1 infected populations. Especially the NRTI-related M184I/V variants, which are among the most frequently encountered mutations in treated patients, are underrepresented in the antiretroviral naïve population. M184I/V mutations are known to have a profound effect on viral replication and tend to revert over time in the new host. However it is debated whether a diminished transmission efficacy of HIV variants with a reduced replication capacity can also contribute to the observed discrepancy in genotypic patterns.As dendritic cells (DCs) play a pivotal role in HIV-1 transmission, we used a model containing primary human Langerhans cells (LCs) and DCs to compare the transmission efficacy M184 variants (HIV-M184V/I/T) to HIV wild type (HIV-WT). As control, we used HIV harboring the NNRTI mutation K103N (HIV-K103N) which has a minor effect on replication and is found at a similar prevalence in treated and untreated individuals.Results
In comparison to HIV-WT, the HIV-M184 variants were less efficiently transmitted to CCR5+ Jurkat T cells by both LCs and DCs. The transmission rate of HIV-K103N was slightly reduced to HIV-WT in LCs and even higher than HIV-WT in DCs. Replication experiments in CCR5+ Jurkat T cells revealed no apparent differences in replication capacity between the mutant viruses and HIV-WT. However, viral replication in LCs and DCs was in concordance with the transmission results; replication by the HIV-M184 variants was lower than replication by HIV-WT, and the level of replication of HIV-K103N was intermediate for LCs and higher than HIV-WT for DCs.Conclusions
Our data demonstrate that drug resistant M184-variants display a reduced replication capacity in LCs and DCs which directly impairs their transmission efficacy. As such, diminished transmission efficacy may contribute to the lower prevalence of drug resistant variants in therapy naive individuals.28.
Schistosomiasis vector snails are subjected to extreme seasonal changes, particularly in ephemeral rivers and lentic waterbodies. In the tropics, aestivation is one of the adaptive strategies for survival and is used by snails in times of extremely high temperatures and desiccation. Aestivation therefore plays an important role in maintaining the transmission of schistosomiasis. This review assesses the possible impacts of climate change on the temporal and spatial distribution of schistosomiasis-transmitting snails with special emphasis on aestivation, and discusses the effect of schistosome infection on aestivation ability. The impacts of parasite development on snails, as well as physiological changes, are discussed with reference to schistosomiasis transmission. This review shows that schistosome-infected snails have lower survival rates during aestivation, and that those that survive manage to get rid of the infection. In general, snail aestivation ability is poor and survival chances diminish with time. Longer dry periods result in fewer, as well as uninfected, snails. However, the ability of the surviving snails to repopulate the habitats is high. 相似文献
29.
L-gulono-γ-lactone oxidase (Gulo) catalyzes the last step of ascorbic acid biosynthesis, which occurs in the kidney of elasmobranchs. This study aimed to clone and sequence gulonolactone oxidase (gulo) from the kidney of the euryhaline freshwater stingray, Himantura signifer, and to determine the effects of acclimation from freshwater to brackish water (salinity 20) on its renal gulo mRNA expression and Gulo activity. We also examined the effects of brackish water acclimation on concentrations of ascorbate, dehydroascorbate and ascorbate + dehydroascorbate in the kidney, brain and gill. The complete cDNA coding sequence of gulo from the kidney of H. signifer contained 1323 bp coding for 440 amino acids. The expression of gulo was kidney-specific, and renal gulo expression decreased significantly by 67% and 50% in fish acclimated to brackish water for 1 day and 6 days, respectively. There was also a significant decrease in renal Gulo activity after 6 days of acclimation to brackish water. Hence, brackish water acclimation led to a decrease in the ascorbic acid synthetic capacity in the kidney of H. signifer. However, there were significant increases in concentrations of ascorbate and ascorbate + dehydroascorbate in the gills (after 1 or 6 days), and a significant increase in the concentration of ascorbate and a significant decrease in the concentration of dehydroascorbate in the brain (after 1 day) of fish acclimated to brackish water. Taken together, our results indicate that H. signifer might experience greater salinity-induced oxidative stress in freshwater than in brackish water, possibly related to its short history of freshwater invasion. These results also suggest for the first time a possible relationship between the successful invasion of the freshwater environment by some euryhaline marine elasmobranchs and the ability of these elasmobranchs to increase the capacity of ascorbic acid synthesis in response to hyposalinity stress. 相似文献
30.