16 research outputs found
Az ischaemiás agykárosodásban szerepet játszó TrpM2 kationcsatorna szerkezet-funkció vizsgálata = Structure-function studies of the TrpM2 cation channel involved in ischaemic brain damage.
MegállapĂtottuk, hogy a csatornát 4 Ca2+ ion kötĹ‘dĂ©se aktiválja. Az aktiváciĂł a Monod-Wymann-Changeux mechanizmust követi, 1 Ca2+ kötĹ‘dĂ©se ~33-szorosra, a 4 ion összesen ~10^6-szorosra, növeli a nyitott-csukott egyensĂşlyi állandĂłt. A Ca2+ kötĹ‘helyei a kaputĂłl intracelluláris irányban találhatĂłk egy vĂ©dett ĂĽregben, közel a pĂłrus nyĂlásához. A nyitott pĂłruson beáramlĂł Ca2+ telĂtĂ©sben tartja az aktiválĂł helyeket, ezĂ©rt intakt sejtekben, ADPR jelenlĂ©tĂ©ben, egy rövid Ca2+ szignál is elnyĂşjtott TRPM2 aktivitást válthat ki. MegállapĂtottuk, hogy az ADPR nagy affinitással (K1/2=1uM) aktivál, de az ADPR-hidrolĂzisnek a csatorna csukĂłdásában játszott szerepĂ©t nem tudtuk tisztázni. Az ADPRázok konzervált ""Nudix-box"" motĂvuma (REFXEE) a TRPM2 NUDT9-H domĂ©njĂ©ben atĂpusos (RILRQE). A NUDT9 enzimben az EF->IL mutáciĂł 1%-ára csökkenti, az EE->KK mutáciĂł felfĂĽggeszti az ADPRáz aktivitást. LĂ©trehoztunk egy ""inaktĂv"" QE->KK Ă©s egy ""hiperaktĂv"" IL->EF TRPM2 mutánst, de e 4 egyedi Ă©s 2 dupla mutáciĂł egyike sem befolyásolta az ADPR iránti affinitást illetve a csukĂłdási sebessĂ©get. Intakt sejtekben az AMP gátolta, mĂg a H2O2, a ciklikus ADPR (cADPR), Ă©s a nikotinsav-adenin-dinukleotid-foszfát (NAADP) aktiválták a TRPM2-t, Ă©s fokozták ADPR iránti Ă©rzĂ©kenysĂ©gĂ©t. Izolált membrán patch-ben megállapĂtottuk, hogy a H2O2, az AMP, Ă©s a cADPR közvetlenĂĽl nem hatnak a TRPM2-re, mĂg az NAADP Ă©s az NAAD kis affinitásĂş parciális agonisták. Tehát intakt sejtekben e modulátorok hatásai közvetettek. | We have revealed that the channel is activated by binding of 4 Ca2+ ions, following the Monod-Wymann-Changeux mechanism. Binding of 1 Ca2+ increases the closed-open equilibrium constant by ~33-fold, the 4 ions altogether by ~10^6-fold. The Ca2+ binding sites are found intracellularly of the gate, in a protected crevice, near the pore entrance, and are kept saturated by Ca2+ flowing through the open pore. Thus, in intact cells, in the presence of ADPR, a single brief Ca2+ spark can elicit prolonged TRPM2 channel activity. We have shown that ADPR activates the channel with high affinity (K1/2=1 uM), but could not clarify the role of ADPR hydrolysis in channel closure. The conserved ADPRase ""Nudix-box"" motif (REFXEE) is atypical in the NUDT9-H domain of TRPM2 (RILRQE). The EF->IL mutation decreases ADPRase activity of the NUDT9 enzyme to ~1%, while the EE->KK mutation completely abolishes it. We constructed an ""inactive"" QE?KK and a ""hiperactive"" IL->EF TRPM2 mutant, but neither of the 4 single and 2 double mutants affected ADPR affinity or channel closing rate. In intact cells AMP inhibits, while H2O2, cyclic ADPR (cADPR), and nicotinic acid-adenin-dinucleotide-phosphate (NAADP) activate TRPM2, and enhance its sensitivity to ADPR. We have found that direct application of H2O2, AMP, and cADPR in isolated patches does not affect the channels, while NAADP and NAAD are low-affinity partial agonists. Thus, in intact cells the effects of these modulators are indirect
On Climate Change, Hydrological Extremes and Water Security in a Globalized World = KlĂmaváltozás, hidrolĂłgiai szĂ©lsĹ‘sĂ©gek Ă©s vĂzbiztonság a (egy) globalizált világban
Insula Insolita – Szigetköz és Bős-Nagymaros párhuzamos története
A közlemĂ©ny cĂ©lja a szigetközi ártĂ©r Ă©s a BĹ‘s-Nagymaros VĂzlĂ©pcsĹ‘rendszer (BNV) párhuzamos törtĂ©netĂ©nek bemutatása Ă©s a törtĂ©net tanulságainak levonása. A BNV-t megelĹ‘zĹ‘ idĹ‘k árvĂzvĂ©delmi Ă©s folyĂłszabályozási munkáinak mellĂ©khatásakĂ©nt, továbbá a Duna fokozott szennyezĹ‘anyag terhelĂ©se miatt, sĂşlyos degradáciĂłs folyamatok indultak be a Szigetközben, melyek megállĂtása Ă©s visszafordĂtása a BNV egyik fontos cĂ©lkitűzĂ©se volt. A mĂşlt század nyolcvanas Ă©veiben kialakult mozgalmak a környezetvĂ©delem jelszavát hirdetve, ám hamis Ă©rvekre hivatkozva a BNV megvalĂłsĂtásának megakadályozását tűztĂ©k ki cĂ©lul. Ezek a mozgalmak, köszönhetĹ‘en a rendszerváltás idejĂ©re kiĂ©pĂtett politikai befolyásuknak, vĂ©gĂĽl elĂ©rtĂ©k a cĂ©ljukat: a BNV beruházását magyar rĂ©szrĹ‘l egyoldalĂşan felfĂĽggesztettĂ©k, majd vĂ©gleg leállĂtották. Szlovákia azonban, mint a nemzetközi beruházás másik rĂ©sztvevĹ‘je, nem adta fel a projektet Ă©s a magyar fĂ©l kiszállásábĂłl adĂłdĂł kĂ©nyszerhelyzethez alkalmazkodva, egyoldalĂşan kivitelezte azt. Ennek következmĂ©nye lett a szigetközi Duna szakasz 30 Ă©vvel ezelĹ‘tti elterelĂ©se. A BNV magyar rĂ©szrĹ‘l törtĂ©nĹ‘ leállĂtása sĂşlyos károkat okozott Magyarországnak: az anyagi, szellemi Ă©s erkölcsi károkozáson tĂşl, az ország de facto elvesztette a beruházás tárgyában Szlovákia ellen indĂtott nemzetközi pert Ă©s mĂ©g az elkĂ©szĂĽlt mű által megtermelt megĂşjulĂł energiábĂłl sem rĂ©szesedik. Az elmĂşlt Ă©vtizedek során vĂ©gzett monitoring tevĂ©kenysĂ©g egyĂ©rtelműen cáfolta a vĂzlĂ©pcsĹ‘ellenes mozgalom ökolĂłgiai katasztrĂłfát vizionálĂł álláspontját. A szigetközi hullámtĂ©r talaj- Ă©s felszĂni vizeinek minĹ‘sĂ©gi Ă©s mennyisĂ©gi viszonyaiban degradáciĂł helyett javulás törtĂ©nt ‒ mĂ©g a BNV elĹ‘tti idĹ‘khöz kĂ©pest is. Ez a javulás a Duna vĂzminĹ‘sĂ©gĂ©ben beállt pozitĂv változásoknak, valamint a Szigetközben kiĂ©pĂĽlt vĂzpĂłtlĂł-rendszernek köszönhetĹ‘. A vĂzpĂłtlĂł-rendszer kiĂ©pĂtĂ©sĂ©vel a magyar vĂzĂĽgyi szolgálat Ăşjfent bebizonyĂtotta, hogy kĂ©pes nagyszerű teljesĂtmĂ©nyek elĂ©rĂ©sĂ©re
Structure of a TRPM2 channel in complex with Ca2+ explains unique gating regulation
Transient receptor potential melastatin 2 (TRPM2) is a Ca2+-permeable cation channel
required for immune cell activation, insulin secretion, and body heat control. TRPM2 is activated by
cytosolic Ca2+, phosphatidyl-inositol-4,5-bisphosphate and ADP ribose. Here, we present the ~3 A°
resolution electron cryo-microscopic structure of TRPM2 from Nematostella vectensis, 63% similar
in sequence to human TRPM2, in the Ca2+-bound closed state. Compared to other TRPM channels,
TRPM2 exhibits unique structural features that correlate with its function. The pore is larger and
more negatively charged, consistent with its high Ca2+ selectivity and larger conductance. The
intracellular Ca2+ binding sites are connected to the pore and cytosol, explaining the unusual
dependence of TRPM2 activity on intra- and extracellular Ca2+. In addition, the absence of a post-
filter motif is likely the cause of the rapid inactivation of human TRPM2. Together, our cryo-EM and
electrophysiology studies provide a molecular understanding of the unique gating mechanism of
TRPM2
The relevance of neck linker docking in the motility of kinesin
Conventional kinesin is a motor protein, which is able to walk along a
microtubule processively. The exact mechanism of the stepping motion and force
generation of kinesin is still far from clear. In this paper we argue that neck
linker docking is a crucial element of this mechanism, without which the
experimentally observed dwell times of the steps could not be explained under a
wide range of loading forces. We also show that the experimental data impose
very strict constraints on the lengths of both the neck linker and its docking
section, which are compatible with the known structure of kinesin.Comment: Accepted for publication in BioSystems as part of the proceedings of
BIOCOMP 200
Neck linker docking coordinates the kinetics of kinesin's heads
Conventional kinesin is a two-headed homodimeric motor protein, which is able
to walk along microtubules processively by hydrolyzing ATP. Its neck linkers,
which connect the two motor domains and can undergo a docking/undocking
transition, are widely believed to play the key role in the coordination of the
chemical cycles of the two motor domains and, consequently, in force production
and directional stepping. Although many experiments, often complemented with
partial kinetic modeling of specific pathways, support this idea, the ultimate
test of the viability of this hypothesis requires the construction of a
complete kinetic model. Considering the two neck linkers as entropic springs
that are allowed to dock to their head domains and incorporating only the few
most relevant kinetic and structural properties of the individual heads, here
we develop the first detailed, thermodynamically consistent model of kinesin
that can (i) explain the cooperation of the heads (including their gating
mechanisms) during walking and (ii) reproduce much of the available
experimental data (speed, dwell time distribution, randomness, processivity,
hydrolysis rate, etc.) under a wide range of conditions (nucleotide
concentrations, loading force, neck linker length and composition, etc.).
Besides revealing the mechanism by which kinesin operates, our model also makes
it possible to look into the experimentally inaccessible details of the
mechanochemical cycle and predict how certain changes in the protein affect its
motion.Comment: to appear in the Biophysical Journa
Application of Electronic Tongue to Soya Drink Discrimination
The objective of the research was to compare the taste attributes of different commercial soya drinks. Furthermore, the task was to determine the effect of different ingredients and processing technologies on the taste attributes of the product. Based on the results of electronic tongue measurements the instrument is able to determine the effect of the applied technology and to distinguish soya juice samples according to sensory preferences. Canonical discriminant analysis showed that the groups of two measurements of the same products were overlapping. Therefore, the electronic tongue measurements are supposed to be of acceptable repeatability. The canonical discriminant analysis showed that the taste attributes of soya juice made of hulled soybeans was beneficial for the taste attributes relative to that of the juice made of not-hulled soybeans. Three main groups could be observed from the analyzed six commercial soya drink samples based on canonical discriminant analysis. There is a group of top market brands having definite taste improver additives and another one containing three products having low amount of additives. However, the group of samples made of soybean and rice is located between the above-mentioned two groups in the discriminant score plot
A Novel Immune-Related Gene Prognostic Index (IRGPI) in Pancreatic Adenocarcinoma (PAAD) and Its Implications in the Tumor Microenvironment
Purpose: Pancreatic adenocarcinoma (PAAD) is one of the most lethal malignancies, with less than 10% of patients surviving more than 5 years. Existing biomarkers for reliable survival rate prediction need to be enhanced. As a result, the objective of this study was to create a novel immune-related gene prognostic index (IRGPI) for estimating overall survival (OS) and to analyze the molecular subtypes based on this index. Materials and procedures: RNA sequencing and clinical data were retrieved from publicly available sources and analyzed using several R software packages. A unique IRGPI and optimum risk model were developed using a machine learning algorithm. The prediction capability of our model was then compared to that of previously proposed models. A correlation study was also conducted between the immunological tumor microenvironment, risk groups, and IRGPI genes. Furthermore, we classified PAAD into different molecular subtypes based on the expression of IRGPI genes and investigated their features in tumor immunology using the K-means clustering technique. Results: A 12-gene IRGPI (FYN, MET, LRSAM1, PSPN, ERAP2, S100A1, IL20RB, MAP3K14, SEMA6C, PRKCG, CXCL11, and GH1) was established, and verified along with a risk model. OS prediction by our model outperformed previous gene signatures. According to the findings of our correlation studies, different risk groups and IRGPI genes were found to be tightly related to tumor microenvironments, and PAAD could be further subdivided into immunologically distinct molecular subtypes based on the expression of IRGPI genes. Conclusion: The current study constructed and verified a unique IRGPI. Furthermore, our findings revealed a connection between the IRGPI and the immunological microenvironment of tumors. PAAD was differentiated into several molecular subtypes that might react differently to immunotherapy. These findings could provide new insights for precision and translational medicine for more innovative immunotherapy strategies
Application of Electronic Tongue for Distinguishing Coffee Samples and Predicting Sensory Attributes
Efforts have been made to predict the sensory profile of coffee samples by instrumental measurement results. The objective of the work was to evaluate the most important sensory attributes of coffee samples prepared from ground roasted coffee by electronic tongue and by sensory panel. Further aim was to predict the Arabica concentration and the main sensory attributes of the different coffee blends by electronic tongue and to analyze the sensitivity of the electronic tongue to the detection of poor quality coffee samples. Five coffee blends with known Arabica and Robusta concentration ratio, five commercially available coffee blends and a poor quality coffee were analyzed. The electronic tongue distinguished the coffee samples according to the Arabica and Robusta content. The sensory panel was able to discriminate the samples based on global aroma, bitterness and coffee aroma intensity (p < 0.01). The Arabica concentration was predicted from the electronic tongue results by PLS with close correlation and low prediction error. Models were developed to predict sensory attributes of the tested coffee samples from the results obtained by the electronic instrument