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1.
Here we address three misconceptions stated by Rice et al. in their observations of our article Paz-y-Mi?o and Espinosa (Evo
Edu Outreach 2:655–675, 2009), published in this journal. The five authors titled their note “The Theory of Evolution is Not an Explanation for the Origin of Life.” First, we argue that it is fallacious to believe that because the formulation of the theory of evolution, as conceived
in the 1800s, did not include an explanation for the origin of life, nor of the universe, the concept of evolution would not
allow us to hypothesize the possible beginnings of life and its connections to the cosmos. Not only Stanley Miller’s experiments
of 1953 led scientists to envision a continuum from the inorganic world to the origin and diversification of life, but also
Darwin’s own writings of 1871. Second, to dismiss the notion of Rice et al. that evolution does not provide explanations concerning
the universe or the cosmos, we identify compelling scientific discussions on the topics: Zaikowski et al. (Evo Edu Outreach
1:65–73, 2008), Krauss (Evo Edu Outreach 3:193–197, 2010), Peretó et al. (Orig Life Evol Biosph 39:395–406, 2009) and Follmann and Brownson (Naturwissenschaften 96:1265–1292, 2009). Third, although we acknowledge that the term Darwinism may not be inclusive of all new discoveries in evolution, and also that creationists and Intelligent Designers hijack the
term to portray evolution as ideology, we demonstrate that there is no statistical evidence suggesting that the word Darwinism
interferes with public acceptance of evolution, nor does the inclusion of the origin of life or the universe within the concept
of evolution. We examine the epistemological and empirical distinction between the theory of evolution and the concept of evolution and conclude that, although the distinction is important, it should not compromise scientific logic. 相似文献
2.
Gilmanov IR Samigullin DV Vyskocil F Nikolsky EE Bukharaeva EA 《Journal of computational neuroscience》2008,25(2):296-307
The local calcium concentration in the active zone of secretion determines the number and kinetics of neurotransmitter quanta
released after the arrival of a nerve action potential in chemical synapses. The small size of mammalian neuromuscular junctions
does not allow direct measurement of the correlation between calcium influx, the state of endogenous calcium buffers determining
the local concentration of calcium and the time course of quanta exocytosis. In this work, we used computer modeling of quanta
release kinetics with various levels of calcium influx and in the presence of endogenous calcium buffers with varying mobilities.
The results of this modeling revealed the desynchronization of quanta release under low calcium influx in the presence of
an endogenous fixed calcium buffer, with a diffusion coefficient much smaller than that of free Ca2+, and synchronization occurred upon adding a mobile buffer. This corresponds to changes in secretion time course parameters
found experimentally (Samigullin et al., Physiol Res 54:129–132, 2005; Bukharaeva et al., J Neurochem 100:939–949, 2007). 相似文献
3.
Jurgen F. Doreleijers Wim F. Vranken Christopher Schulte Jundong Lin Jonathan R. Wedell Christopher J. Penkett Geerten W. Vuister Gert Vriend John L. Markley Eldon L. Ulrich 《Journal of biomolecular NMR》2009,45(4):389-396
Several pilot experiments have indicated that improvements in older NMR structures can be expected by applying modern software
and new protocols (Nabuurs et al. in Proteins 55:483–186, 2004; Nederveen et al. in Proteins 59:662–672, 2005; Saccenti and Rosato in J Biomol NMR 40:251–261, 2008). A recent large scale X-ray study also has shown that modern software can significantly improve the quality of X-ray structures
that were deposited more than a few years ago (Joosten et al. in J. Appl Crystallogr 42:376–384, 2009; Sanderson in Nature 459:1038–1039, 2009). Recalculation of three-dimensional coordinates requires that the original experimental data are available and complete,
and are semantically and syntactically correct, or are at least correct enough to be reconstructed. For multiple reasons,
including a lack of standards, the heterogeneity of the experimental data and the many NMR experiment types, it has not been
practical to parse a large proportion of the originally deposited NMR experimental data files related to protein NMR structures.
This has made impractical the automatic recalculation, and thus improvement, of the three dimensional coordinates of these
structures. We here describe a large-scale international collaborative effort to make all deposited experimental NMR data
semantically and syntactically homogeneous, and thus useful for further research. A total of 4,014 out of 5,266 entries were
‘cleaned’ in this process. For 1,387 entries, human intervention was needed. Continuous efforts in automating the parsing
of both old, and newly deposited files is steadily decreasing this fraction. The cleaned data files are available from the
NMR restraints grid at . 相似文献
4.
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6.
Meiotic prophase serves as an arena for the interplay of two important cellular activities, meiotic recombination and synapsis of homologous chromosomes. Synapsis is mediated by the synaptonemal complex (SC), originally characterized as a structure linked to pairing of meiotic chromosomes (Moses (1958) J Biophys Biochem Cytol 4:633–638). In 1975, the first electron micrographs of human pachytene stage SCs were presented (Moses et al. (1975) Science 187:363–365) and over the next 15 years the importance of the SC to normal meiotic progression in human males and females was established (Jhanwar and Chaganti (1980) Hum Genet 54:405–408; Pathak and Elder (1980) Hum Genet 54:171–175; Solari (1980) Chromosoma 81:315–337; Speed (1984) Hum Genet 66:176–180; Wallace and Hulten (1985) Ann Hum Genet 49(Pt 3):215–226). Further, these studies made it clear that abnormalities in the assembly or maintenance of the SC were an important contributor to human infertility (Chaganti et al. (1980) Am J Hum Genet 32:833–848; Vidal et al. (1982) Hum Genet 60:301–304; Bojko (1983) Carlsberg Res Commun 48:285–305; Bojko (1985) Carlsberg Res Commun 50:43–72; Templado et al. (1984) Hum Genet 67:162–165; Navarro et al. (1986) Hum Reprod 1:523–527; Garcia et al. (1989) Hum Genet 2:147–53). However, the utility of these early studies was limited by lack of information on the structural composition of the SC and the identity of other SC-associated proteins. Fortunately, studies of the past 15 years have gone a long way toward remedying this problem. In this minireview, we highlight the most important of these advances as they pertain to human meiosis, focusing on temporal aspects of SC assembly, the relationship between the SC and meiotic recombination, and the contribution of SC abnormalities to human infertility.The synaptonemal complex–50 years 相似文献
7.
Temperature controls a latitudinal gradient in the proportion of watershed nitrogen exported to coastal ecosystems 总被引:5,自引:4,他引:1
Increased export of biologically available nitrogen (N) to the coastal zone is strongly linked to eutrophication, which is
a major problem in coastal marine ecosystems (NRC (2000) Clean Coastal Waters: Understanding and Reducing the Effects of Nutrient Pollution. National Academy Press, Washington,
DC; Bricker et al. (1999) National Estuarine Eutrophication Assessment. Effects of nutrient enrichment in the nation’s estuaries. NOAA-NOS Special
Projects Office, Silver Spring, MD). However, not all of the nitrogen input to a watershed is exported to the coast (Howarth
et al. (1996) Biogeochemistry 35:75–139; Jordan and Weller (1996) Bioscience 46:655–664). Global estimates of nitrogen export to coasts have been taken to be 25% of watershed input, based
largely on northeastern U.S. observations (Galloway et al. (2004) Biogeochemistry 70:153–226; Boyer et al. (2006) Global Biogeochem Cycle 20:Art. No. GB1S91). We applied the N budgeting methodology developed for the International SCOPE
Nitrogen project (Howarth et al. (1996) Biogeochemistry 35:75–139; Boyer et al. (2002) Biogeochemistry 57:137–169) to 12 watersheds in the southeastern U.S., and compared them with estimates of N export for
16 watersheds in the northeastern U.S. (Boyer et al. (2002) Biogeochemistry 57:137–169). In southeastern watersheds, average N export was only 9% of input, suggesting the need for
downward revision of global estimates. The difference between northern and southern watersheds is not a function of the absolute
value of N inputs, which spanned a comparable range and were positively related to export in both cases. Rather, the proportion
of N exported was significantly related to average watershed temperature (% N export = 58.41 e−0.11 * temperature; R
2 = 0.76), with lower proportionate nitrogen export in warmer watersheds. In addition, we identified a threshold in proportionate
N export at 38°N latitude that corresponds to a reported breakpoint in the rate of denitrification at 10–12°C. We hypothesize
that temperature, by regulating denitrification, results in increased proportionate N export at higher latitudes. Regardless
of the mechanism, these observations suggest that temperature increases associated with future climate change may well reduce
the amount of nitrogen that reaches estuaries, which will have implications for coastal eutrophication. 相似文献
8.
Under intracellular recording, we studied the effect of ATP on nerve cells of the rat intact nodose ganglion. The resting
membrane potential of the examined neurons was, on average, –60.3 ± 1.4 mV (n = 84); among such units, 88% were classified as C cells. Local application of 2 mM ATP to the surface of the ganglion using
a modified laminar flow system led to depolarization of neurons by 7.1 ± 0.9 mV, on average (n = 19). A blocker of P2X receptors, PPADS (100 μM), suppressed these depolarization responses, decreasing their amplitude,
on average, to 16 ± 3% (n = 3) of the initial value. The obtained data indicate that an overwhelming majority of neurons of the intact nodose ganglion
possess functional P2X receptors on their membranes. The absence of the corresponding responses in a considerable part of
neurons of intact spinal ganglia [13-15] was, apparently, determined by the fact that P2X receptors in the course of the described experiments had enough time to
desensitize before ATP reached the effective concentration. 相似文献
9.
Etzel CJ Chen WV Shepard N Jawaheer D Cornelis F Seldin MF Gregersen PK Amos CI;North American Rheumatoid Arthritis Consortium 《Human genetics》2006,119(6):634-641
Meta-analysis is being increasingly used as a tool for integrating data from different studies of complex phenotypes, because the power of any one study to identify causal loci is limited. We applied a novel meta-analytical approach (Loesgen et al. in Genet Epidemiol 21(Suppl 1):S142–S147, 2001) in compiling results from four studies of rheumatoid arthritis in Caucasians including two studies from NARAC (Jawaheer et al. in Am J Hum Genet 68:927–936, 2001; Jawaheer et al. in Arthritis Rheum 48:906–916, 2003), one study from the UK (MacKay et al. in Arthritis Rheum 46:632–639, 2001) and one from France (Cornelis et al. in Proc Natl Acad Sci USA 95:10746–10750, 1998). For each study, we obtained NPL scores by performing interval mapping (2 cM intervals) using GeneHunter2 (Kruglyak et al. in Am J Hum Genet 58:1347–1363, 1996; Markianos et al. in Am J Hum Genet 68:963–977, 2001). The marker maps differed among the three consortium groups, therefore, the marker maps were aligned after the interval mapping was completed and the NPL scores that were within 1 cM of each other were combined using the method of Loesgen et al. (Genet Epidemiol 21(Suppl 1):S142–S147, 2001) by calculating the weighted average of the NPL score. This approach avoids some problems in analysis encountered by using GeneHunter2 when some markers in the sample are not genotyped. This procedure provided marginal evidence (P<0.05) of linkage on chromosome 1, 2, 5 and 18, strong evidence (P<0.01) on chromosomes 8 and 16, and overwhelming evidence in the HLA region of chromosome 6. 相似文献
10.
A Closer Look at Amphetamine-Induced Reverse Transport and Trafficking of the Dopamine and Norepinephrine Transporters 总被引:1,自引:0,他引:1
Amphetamine (AMPH) and its derivatives are regularly used in the treatment of a wide array of disorders such as attention-deficit
hyperactivity disorder (ADHD), obesity, traumatic brain injury, and narcolepsy (Prog Neurobiol 75:406–433, 2005; J Am Med Assoc 105:2051–2054, 1935; J Am Acad Child Adolesc Psychiatry 41:514–521, 2002; Neuron 43:261–269, 2004; Annu Rev Pharmacol Toxicol 47:681–698, 2007; Drugs Aging 21:67–79, 2004). Despite the important medicinal role for AMPH, it is more widely known for its psychostimulant and addictive properties
as a drug of abuse. The primary molecular targets of AMPH are both the vesicular monoamine transporters (VMATs) and plasma
membrane monoamine—dopamine (DA), norepinephrine (NE), and serotonin (5-HT)—transporters. The rewarding and addicting properties
of AMPH rely on its ability to act as a substrate for these transporters and ultimately increase extracellular levels of monoamines.
AMPH achieves this elevation in extracellular levels of neurotransmitter by inducing synaptic vesicle depletion, which increases
intracellular monoamine levels, and also by promoting reverse transport (efflux) through plasma membrane monoamine transporters
(J Biol Chem 237:2311–2317, 1962; Med Exp Int J Exp Med 6:47–53, 1962; Neuron 19:1271–1283, 1997; J Physiol 144:314–336, 1958; J Neurosci 18:1979–1986, 1998; Science 237:1219–1223, 1987; J Neurosc 15:4102–4108, 1995). This review will focus on two important aspects of AMPH-induced regulation of the plasma membrane monoamine transporters—transporter
mediated monoamine efflux and transporter trafficking. 相似文献
11.
Pectin is one of the major cell wall polysaccharides found in dicotyledonous plants. We have solubilized and partially purified
a β-(1→4)-galactosyltransferase (GalT) involved in the synthesis of the β-(1→4)-galactan side chains of pectin. The enzyme
protein was almost completely solubilized by mixing a crude microsomal preparation of etiolated 6-day-old soybean (Glycine max Merr.) hypocotyls with a detergent, Triton X-100 (0.75%, w/v), in buffer. The solubilized enzyme was partially purified by
ion-exchange chromatography. The crude membrane-bound GalT transferred Gal from UDP-Gal onto 2-aminobenzamide (AB)-derivatized
β-(1→4)-galactoheptaose (Gal7-AB), leading to the formation of Gal8–11-AB by attachment of a series of one to four galactosyl residues; this is similar to what has previously been observed for
2-aminopyridine-derivatized β-(1→4)-galactooligomer acceptors (Konishi et al. in Planta 218:833–842, 2004). The partially purified GalT, by contrast, was able to transfer more than 25 galactosyl residues and elongated the chains
to about Gal35-AB, thus almost reaching the length (43–47 Gal units) of native β-(1→4)-galactan side chains found in pectic polysaccharides
from soybean cotyledons (Nakamura et al. in Biosci Biotechnol Biochem 66:1301–1313, 2002). Enzyme activity increased with increasing chain length of β-(1→4)-galactooligomers and reached maximal activity at heptaose,
whereas galactooligomers higher than heptaose showed lower acceptor efficiency.
Sugars described in this paper belong to the d-series unless otherwise noted. 相似文献
12.
Roger Bill 《Dialectical Anthropology》2010,34(3):395-417
Jack Kerouac, the author of On The Road, was a central figure of the Beat Generation, a generation which rebelled against middle-class conformity in post–World War
II America. Kerouac described himself as “a religious wanderer” (Kerouac 2006: 2), but an examination of his texts and life suggest his travels may also be understood as tourism. Viewed through the prism
of tourism, this study will argue, for example, that MacCannell’s notion of the tourist’s quest for reality and authenticity
(MacCannell 1989: 3) provides some insight into why Kerouac wrote that just south of Macon, Georgia, he and his travelling companion Neal
Cassady stopped and got out of the car, “and suddenly both of us were stoned with joy to realize that in the darkness all
around us was fragrant green grass and the smell of fresh manure and warm waters” (Kerouac 1957: 115). As Kerouac rebelled against being, as one of his protagonists in The Dharma Bums put it, “imprisoned in a system of work, produce, consume, work, produce, consume” (Kerouac 2006: 73) he travelled across America on a rapidly improving network of highways, turning “mobility into a retreat” (Holladay
and Holton 2009: 42). Kerouac alternately identified himself as a hobo (Kerouac 1973: 181) and “not a real hobo” (Kerouac 1973: 173), but this article asks whether Kerouac’s travels were those of the last in a line of wanderers rebelling against conformity
and modernization or a precursor of mobile mass tourism in America. 相似文献
13.
Siderophore production by marine-derived fungi 总被引:1,自引:0,他引:1
14.
Christina M. Grozinger Gene E. Robinson 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》2007,193(4):461-470
Pheromones cause dramatic changes in behavior and physiology, and are critical for honey bee colony organization. Queen mandibular
pheromone (QMP) regulates multiple behaviors in worker bees (Slessor et al. in J Chem Ecol 31(11):2731–2745, 2005). We also identified genes whose brain expression levels were altered by exposure to QMP (Grozinger et al. in Proc Natl Acad
Sci USA 100(Suppl 2):14519–14525, 2003). Krüppel-homolog 1 (Kr-h1) RNA levels were significantly downregulated by QMP, and were higher in foragers than in nurses (Whitfield et al. in Science
302(5643):296–299, 2003). Here we report on results of behavioral and pharmacological experiments that characterize factors regulating expression
of Kr-h1. Foragers have higher brain levels of Kr-h1 than in-hive bees, regardless of age and pheromone exposure. Furthermore, forager Kr-h1 levels were not affected by QMP. Since the onset of foraging is caused, in part, by increasing juvenile hormone blood titers
and brain octopamine levels, we investigated the effects of octopamine and methoprene (a juvenile hormone analog) on Kr-h1 expression. Methoprene produced a marginal (not significant) increase in Kr-h1 expression, but Kr-h1 brain levels in methoprene-treated bees were no longer downregulated by QMP. Octopamine did not modulate Kr-h1 expression. Our results demonstrate that the gene expression response to QMP is not hard-wired in the brain but is instead
dependent on worker behavioral state. 相似文献
15.
Simulation of biological evolution under attack,but not really: a response to Meester 总被引:1,自引:0,他引:1
The leading Intelligent Design theorist William Dembski (Rowman & Littlefield, Lanham MD, 2002) argued that the first No Free Lunch theorem, first formulated by Wolpert and Macready (IEEE Trans Evol Comput 1: 67–82,
1997), renders Darwinian evolution impossible. In response, Dembski’s critics pointed out that the theorem is irrelevant to biological
evolution. Meester (Biol Phil 24: 461–472, 2009) agrees with this conclusion, but still thinks that the theorem does apply to simulations of evolutionary processes. According
to Meester, the theorem shows that simulations of Darwinian evolution, as these are typically set in advance by the programmer,
are teleological and therefore non-Darwinian. Therefore, Meester argues, they are useless in showing how complex adaptations
arise in the universe. Meester uses the term “teleological” inconsistently, however, and we argue that, no matter how we interpret
the term, a Darwinian algorithm does not become non-Darwinian by simulation. We show that the NFL theorem is entirely irrelevant
to this argument, and conclude that it does not pose a threat to the relevance of simulations of biological evolution. 相似文献
16.
Zachary Cooper Michael Greenwood Borbala Mazzag 《Bulletin of mathematical biology》2009,71(7):1543-1579
We investigate the role of heterogeneous expression of IP3R and RyR in generating diverse elementary Ca2+ signals. It has been shown empirically (Wojcikiewicz and Luo in Mol. Pharmacol. 53(4):656–662, 1998; Newton et al. in J. Biol. Chem. 269(46):28613–28619, 1994; Smedt et al. in Biochem. J. 322(Pt. 2):575–583, 1997) that tissues express various proportions of IP3 and RyR isoforms and this expression is dynamically regulated (Parrington et al. in Dev. Biol. 203(2):451–461, 1998; Fissore et al. in Biol. Reprod. 60(1):49–57, 1999; Tovey et al. in J. Cell Sci. 114(Pt. 22):3979–3989, 2001). Although many previous theoretical studies have investigated the dynamics of localized calcium release sites (Swillens
et al. in Proc. Natl. Acad. Sci. U.S.A. 96(24):13750–13755, 1999; Shuai and Jung in Proc. Natl. Acad. Sci. U.S.A. 100(2):506–510, 2003a; Shuai and Jung in Phys. Rev. E, Stat. Nonlinear Soft Matter Phys. 67(3 Pt. 1):031905, 2003b; Thul and Falcke in Biophys. J. 86(5):2660–2673, 2004; DeRemigio and Smith in Cell Calcium 38(2):73–86, 2005; Nguyen et al. in Bull. Math. Biol. 67(3):393–432, 2005), so far all such studies focused on release sites consisting of identical channel types. We have extended an existing mathematical
model (Nguyen et al. in Bull. Math. Biol. 67(3):393–432, 2005) to release sites with two (or more) receptor types, each with its distinct channel kinetics. Mathematically, the release
site is represented by a transition probability matrix for a collection of nonidentical stochastically gating channels coupled
through a shared Ca2+ domain. We demonstrate that under certain conditions a previously defined mean-field approximation of the coupling strength
does not accurately reproduce the release site dynamics. We develop a novel approximation and establish that its performance
in these instances is superior. We use this mathematical framework to study the effect of heterogeneity in the Ca2+-regulation of two colocalized channel types on the release site dynamics. We consider release sites consisting of channels
with both Ca2+-activation and inactivation (“four-state channels”) and channels with Ca2+-activation only (“two-state channels”) and show that for the appropriate parameter values, synchronous channel openings within
a release site with any proportion of two-state to four-state channels are possible, however, the larger the proportion of
two-state channels, the more sensitive the dynamics are to the exact spatial positioning of the channels and the distance
between channels. Specifically, the clustering of even a small number of two-state channels interferes with puff/spark termination
and increases puff durations or leads to a tonic response. 相似文献
17.
Oktaviani NA Otten R Dijkstra K Scheek RM Thulin E Akke M Mulder FA 《Biomolecular NMR assignments》2011,5(1):79-84
Here we present the 100% complete assignment chemical shift of non-labile 1H, 15N and 13C nuclei of Calbindin D9k P43G. The assignment includes all non-exchangeable side chain nuclei, including ones that are rarely reported, such as LysNζ
as well as the termini. NMR experiments required to achieve truly complete assignments are discussed. To the best of our knowledge
our assignments for Calbindin D9k extend beyond previous studies reaching near-completeness (Vis et al. in Biochem 33:14858–14870, 1994; Yamazaki et al. in J Am Chem Soc 116:6464–6465, 1994; Yamazaki et al. in Biochem 32:5656–5669, 1993b). 相似文献
18.
Agarwal B 《Journal of bioenergetics and biomembranes》2011,43(3):299-310
ATP, the ‘universal biological energy currency’, is synthesized by utilizing energy either from oxidation of fuels or from
light, via the process of oxidative and photo-phosphorylation respectively. The process is mediated by the enzyme F1F0-ATP synthase, using the free energy of ion gradients in the final energy catalyzing step, i.e., the synthesis of ATP from
ADP and inorganic phosphate (Pi). The details of the molecular mechanism of ATP synthesis are among the most important fundamental issues in biology and
hence need to be properly understood. In this work, a role for anions in making ATP has been found. New experimental data
has been reported on the inhibition of ATP synthesis at nanomolar concentrations by the potent, specific anion channel blockers
4,4′-diisothiocyanostilbene-2, 2′-disulphonic acid (DIDS) and tributyltin chloride (TBTCl). Based on these inhibition studies,
attention has been drawn to anion translocation (in addition to proton translocation) as a requirement for ATP synthesis.
The type of inhibition has been quantified and an overall kinetic scheme for mixed inhibition that explains the data has been
evolved. The experimental data and the type of inhibition found have been interpreted in the light of the torsional mechanism
of energy transduction and ATP synthesis (Nath J Bioenerg Biomembr 42:293–300, 2010a; J Bioenerg Biomembr 42:301–309, 2010b). This detailed and unified mechanism resolves long-standing problems and inconsistencies in the first theories (Slater Nature
172:975–978, 1953; Williams J Theor Biol 1:1–17, 1961; Mitchell Nature 191:144–148, 1961; Mitchell Biol Rev 41:445–502, 1966), makes several novel predictions that are experimentally verifiable (Nath Biophys J 90:8–21, 2006a; Process Biochem 41:2218–2235, 2006b), and provides us with a new and fruitful paradigm in bioenergetics. The interpretation presented here provides intelligent
answers to the unexplained existing results in the literature. It is shown that mechanistic interpretation of the experimental
data requires substantial addition to available conceptual foundations such that present concepts, theories, and mechanisms
must be revised. 相似文献
19.
Finn T. O. Krogstad William C. Griffith Eric M. Vigoren Elaine M. Faustman 《Journal of applied phycology》2009,21(6):745-746
Re-examining the reported observation and analysis in Wohlgeshaffen et al. (Journal of Applied Phycology 4(4):297–310, 1992) shows that their observed shellfish domoic acid (DA) concentrations do not support the reported 17% · d−1 depuration rate for blue mussels but instead support depuration rates of 87% · d−1 (fractional) or 200%/day (exponential). This error could have impacts for public health, shellfish management, and other
biological research. This paper identifies a large underestimation in filtration efficiency in Wohlgeshaffen et al. (1992). 相似文献
20.
Koh Takeuchi Gregory Heffron Zhen-Yu J. Sun Dominique P. Frueh Gerhard Wagner 《Journal of biomolecular NMR》2010,47(4):271-282
Heteronuclear direct-detection experiments, which utilize the slower relaxation properties of low γ nuclei, such as 13C have recently been proposed for sequence-specific assignment and structural analyses of large, unstructured, and/or paramagnetic
proteins. Here we present two novel 15N direct-detection experiments. The CAN experiment sequentially connects amide 15N resonances using 13Cα chemical shift matching, and the CON experiment connects the preceding 13C′ nuclei. When starting from the same carbon polarization, the intensities of nitrogen signals detected in the CAN or CON
experiments would be expected four times lower than those of carbon resonances observed in the corresponding 13C-detecting experiment, NCA-DIPAP or NCO-IPAP (Bermel et al. 2006b; Takeuchi et al. 2008). However, the disadvantage due to the lower γ is counteracted by the slower 15N transverse relaxation during detection, the possibility for more efficient decoupling in both dimensions, and relaxation
optimized properties of the pulse sequences. As a result, the median S/N in the 15N observe CAN experiment is 16% higher than in the 13C observe NCA-DIPAP experiment. In addition, significantly higher sensitivity was observed for those residues that are hard
to detect in the NCA-DIPAP experiment, such as Gly, Ser and residues with high-field Cα resonances. Both CAN and CON experiments are able to detect Pro resonances that would not be observed in conventional proton-detected
experiments. In addition, those experiments are free from problems of incomplete deuterium-to-proton back exchange in amide
positions of perdeuterated proteins expressed in D2O. Thus, these features and the superior resolution of 15N-detected experiments provide an attractive alternative for main chain assignments. The experiments are demonstrated with
the small model protein GB1 at conditions simulating a 150 kDa protein, and the 52 kDa glutathione S-transferase dimer, GST. 相似文献