Electrosprayed protein ions can retain native-like conformations. The intramolecular contacts that stabilize these compact gas-phase structures remain poorly understood. Recent work has uncovered ...abundant salt bridges in electrosprayed proteins. Salt bridges are zwitterionic BH+/A– contacts. The low dielectric constant in the vacuum strengthens electrostatic interactions, suggesting that salt bridges could be a key contributor to the retention of compact protein structures. A problem with this assertion is that H+ are mobile, such that H+ transfer can convert salt bridges into neutral B0/HA0 contacts. This possible salt bridge annihilation puts into question the role of zwitterionic motifs in the gas phase, and it calls for a detailed analysis of BH+/A– versus B0/HA0 interactions. Here, we investigate this issue using molecular dynamics (MD) simulations and electrospray experiments. MD data for short model peptides revealed that salt bridges with static H+ have dissociation energies around 700 kJ mol–1. The corresponding B0/HA0 contacts are 1 order of magnitude weaker. When considering the effects of mobile H+, BH+/A– bond energies were found to be between these two extremes, confirming that H+ migration can significantly weaken salt bridges. Next, we examined the protein ubiquitin under collision-induced unfolding (CIU) conditions. CIU simulations were conducted using three different MD models: (i) Positive-only runs with static H+ did not allow for salt bridge formation and produced highly expanded CIU structures. (ii) Zwitterionic runs with static H+ resulted in abundant salt bridges, culminating in much more compact CIU structures. (iii) Mobile H+ simulations allowed for the dynamic formation/annihilation of salt bridges, generating CIU structures intermediate between scenarios (i) and (ii). Our results uncover that mobile H+ limit the stabilizing effects of salt bridges in the gas phase. Failure to consider the effects of mobile H+ in MD simulations will result in unrealistic outcomes under CIU conditions.
Plasma flows related to magnetospheric convection or ULF waves often increase cold ions' kinetic energy sufficiently above the spacecraft potential energy so that these ions can be detected by ...charged particle instrumentation. Other detection methods also reveal the presence of cold ions even in the absence of flows. By applying such methods to complementary data sets obtained from particle, electric field, and magnetic field measurements by the multiple Time History of Events and Macroscale Interactions during Substorms spacecraft, we study the occurrence rates of cold ions and properties—composition, densities, and temperatures—of the dominant species (H+, He+, and O+) near the equatorial magnetosphere. As plasma flows occur at all magnetic local time (MLT) sectors, predominantly outside the plasmasphere, they enable studies of cold ion occurrence and properties as functions of geocentric distance (up to 13 RE) and MLT. Of the dominant cold ions, protons are most abundant on the dayside and coldest almost everywhere. By comparison, the heavier ions show evidence of larger abundance on the nightside and higher temperatures at most locations: median nHe+/nH+ is ≤0.1 in the afternoon, ≥1.0 on the nightside, and near 0.5 at dawn; median nO+/nH+ exceeds 1 throughout the nightside and around dawn at smaller geocentric distances but varies from 0.0 to 0.1 in the afternoon; and TO+ > THe+ everywhere. By isolating the convective particle drift component of measured ion flows, we deduce that nightside/dawn/prenoon cold ions are components of the warm plasma cloak while cold ions near noon/afternoon are likely of mixed origin, including nightside/cusp outflow, plasmaspheric plumes, and the solar wind.
Key Points
Cold ion occurrence and properties are studied near the equatorial magnetosphere
Protons: the most abundant cold ion species on dayside and cooler everywhere
Helium and oxygen ions can exceed proton density and temperature on nightside
A lack of electrical conductivity and structural organization in currently available biomaterial scaffolds limits their utility for generating physiologically representative models of functional ...cardiac tissue. Here we report on the development of scalable, graphene-functionalized topographies with anisotropic electrical conductivity for engineering the structural and functional phenotypes of macroscopic cardiac tissue constructs. Guided by anisotropic electroconductive and topographic cues, the tissue constructs displayed structural property enhancement in myofibrils and sarcomeres, and exhibited significant increases in the expression of cell-cell coupling and calcium handling proteins, as well as in action potential duration and peak calcium release.
It is believed that electrosprayed proteins and protein complexes can retain solution-like conformations in the gas phase. However, the lack of high-resolution structure determination methods for ...gaseous protein ions implies that their properties remain poorly understood. Many practitioners tackle this difficulty by complementing mass spectrometry-based experiments with molecular dynamics (MD) simulations. It is a potential problem that the standard MD force fields used for this purpose (such as OPLS-AA/L and CHARMM) were optimized for solution conditions. The question whether these force fields produce meaningful gas-phase data has received surprisingly little attention. Standard force fields are overpolarized to account for an aqueous environment, i.e., atomic charges and intramolecular dipole moments are ∼20% larger than predicted by gas-phase ab initio methods. Here, we examined the implications of this overpolarization by conducting a series of MD simulations on electrosprayed proteins. Force fields were modified via a charge scaling factor (CSF), while ensuring that the net protein charge remained unchanged. CSF = 0.8 should roughly eliminate water-associated overpolarization. Gas-phase CHARMM simulations on myoglobin with CSF = 0.8 and with unmodified parameters (CSF = 1) yielded similar results, preserving a compact structure that was consistent with ion mobility experiments. Major structural changes caused by weakened charge–dipole and dipole–dipole contacts occurred only when lowering CSF to physically unreasonable values (0.5 and 0.1). Similar results were obtained in mobile-proton OPLS-AA/L simulations on the collision-induced dissociation of transthyretin. Our data support the view that gas-phase MD simulations with standard (solution) force fields are suitable for modeling gaseous protein ions in a semiquantitative manner. Although this is welcome news for the mass spectrometry community, it is hoped that dedicated gas-phase MD force fields will become available in the near future.
Establishment, specification, and validation of synaptic connections are thought to be mediated by interactions between pre- and postsynaptic cell-adhesion molecules. Arguably, the best-characterized ...transsynaptic interactions are formed by presynaptic neurexins, which bind to diverse postsynaptic ligands. In a proteomic screen of neurexin-1 (Nrxn1) complexes immunoisolated from mouse brain, we identified carbonic anhydrase-related proteins CA10 and CA11, two homologous, secreted glycoproteins of unknown function that are predominantly expressed in brain. We found that CA10 directly binds in a cis configuration to a conserved membrane-proximal, extracellular sequence of α- and β-neurexins. The CA10–neurexin complex is stable and stoichiometric, and results in formation of intermolecular disulfide bonds between conserved cysteine residues in neurexins and CA10. CA10 promotes surface expression of α- and β-neurexins, suggesting that CA10 may form a complex with neurexins in the secretory pathway that facilitates surface transport of neurexins. Moreover, we observed that the Nrxn1 gene expresses from an internal 3′ promoter a third isoform, Nrxn1γ, that lacks all Nrxn1 extracellular domains except for the membrane-proximal sequences and that also tightly binds to CA10. Our data expand the understanding of neurexin-based transsynaptic interaction networks by providing further insight into the interactions nucleated by neurexins at the synapse.
Electromagnetic ion cyclotron (EMIC) wave generation and propagation in Earth's magnetosphere depend on readily measurable hot (a few to tens of keV) plasma sheet ions, elusive plasmaspheric or ...ionospheric cold (sub‐eV to a few eV) ions, and partially heated warm ions (tens to hundreds of eV). Previous work has assumed all low‐energy ions are cold and not considered possible effects of warm ions. Using measurements by multiple Time History of Events and Macroscale Interactions during Substorms spacecraft, we analyze four typical EMIC wave events in the four magnetic local time sectors and consider the properties of both cold and warm ions supplied from previous statistical studies to interpret the wave observations using linear theory. As expected, we find that dusk EMIC waves grow due to the presence of drifting hot anisotropic protons and cold plasmaspheric ions with a dominant cold proton component. Near midnight, EMIC waves are less common because warm heavy ions that suppress wave growth are more abundant there. The waves can grow when cold, plume‐like density enhancements are present, however. Dawn EMIC waves, known for their peculiar properties, are generated away from the equator and change polarization during propagation through the warm plasma cloak. Noon EMIC waves can also be generated nonlocally and their properties modified during propagation by a plasmaspheric plume combined with low‐energy ions from solar and terrestrial sources. Accounting for multiple ion species, measured wave dispersion, and propagation characteristics can explain previously elusive EMIC wave properties and are therefore important for future studies of EMIC wave effects on energetic particle depletion.
Key Points
Using linear theory with typical low‐energy composition to explore wave growthAnalyzing how low‐energy components influence wave growth and propagationUsing waves as diagnostic of environmental properties including composition
The increased treatment of metastatic castration-resistant prostate cancer (mCRPC) with second-generation antiandrogen therapies (ADT) has coincided with a greater incidence of lethal, aggressive ...variant prostate cancer (AVPC) tumors that have lost dependence on androgen receptor (AR) signaling. These AR-independent tumors may also transdifferentiate to express neuroendocrine lineage markers and are termed neuroendocrine prostate cancer (NEPC). Recent evidence suggests kinase signaling may be an important driver of NEPC. To identify targetable kinases in NEPC, we performed global phosphoproteomics comparing several AR-independent to AR-dependent prostate cancer cell lines and identified multiple altered signaling pathways, including enrichment of RET kinase activity in the AR-independent cell lines. Clinical NEPC patient samples and NEPC patient-derived xenografts displayed upregulated RET transcript and RET pathway activity. Genetic knockdown or pharmacologic inhibition of RET kinase in multiple mouse and human models of NEPC dramatically reduced tumor growth and decreased cell viability. Our results suggest that targeting RET in NEPC tumors with high RET expression could be an effective treatment option. Currently, there are limited treatment options for patients with aggressive neuroendocrine prostate cancer and none are curative. IMPLICATIONS: Identification of aberrantly expressed RET kinase as a driver of tumor growth in multiple models of NEPC provides a significant rationale for testing the clinical application of RET inhibitors in patients with AVPC.
Although hepatocellular carcinoma (HCC) has become a common indication for liver transplantation (LT), intrahepatic cholangiocarcinoma (ICC) and combined hepatocellular carcinoma–cholangiocarcinoma ...(cHCC‐CCA) are historically contraindicated due to their aggressive behavior. On the basis of recent experiences, some groups have proposed a clinical trial investigating the role of LT for patients with early cholangiocarcinoma (CCA), defined as a single lesion ≤ 2 cm. The purpose of this study is to assess the clinicopathologic features and outcomes following LT for patients who were initially diagnosed with HCC and subsequently found to have either ICC or cHCC‐CCA on explant. Patients with the diagnosis of primary liver cancer (PLC) after LT from a single center were retrospectively reviewed. Outcomes for patients with early CCA were compared with patients with HCC within Milan criteria (MC). Out of 618 patients transplanted with PLC, 44 patients were found to have CCA on explant. On the basis of preoperative imaging, 12 patients met criteria for early CCA and were compared with 319 patients who had HCC within MC. The 1‐ and 5‐year overall survival for early CCA versus HCC was 63.6% versus 90.0% and 63.6% versus 70.3% (log‐rank, P = 0.25), respectively. Overall recurrence was 33.3% for early CCA versus 11% for HCC. On explant the patients with CCA were more likely understaged with higher tumor grade and vascular invasion. In conclusion, patients with CCA present a diagnostic challenge, which often leads to the finding of more aggressive lesions on explant after LT, higher recurrence rates, and worse post‐LT survival. Careful consideration of this diagnostic conundrum needs to be made before a clinical trial is undertaken. Liver Transplantation 24 634–644 2018 AASLD.
Patients with cancer are at increased risk of hospitalisation and mortality following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, the SARS-CoV-2 phenotype ...evolution in patients with cancer since 2020 has not previously been described. We therefore evaluated SARS-CoV-2 on a UK populationscale from 01/11/2020-31/08/2022, assessing case-outcome rates of hospital assessment(s), intensive care admission and mortality. We observed that the SARS-CoV-2 disease phenotype has become less severe in patients with cancer and the non-cancer population. Case-hospitalisation rates for patients with cancer dropped from 30.58% in early 2021 to 7.45% in 2022 while case-mortality rates decreased from 20.53% to 3.25%. However, the risk of hospitalisation and mortality remains 2.10x and 2.54x higher in patients with cancer, respectively. Overall, the SARS-CoV-2 disease phenotype is less severe in 2022 compared to 2020 but patients with cancer remain at higher risk than the non-cancer population. Patients with cancer must therefore be empowered to live more normal lives, to see loved ones and families, while also being safeguarded with expanded measures to reduce the risk of transmission.
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