SummaryAcquired protection from P
lasmodium falciparum malaria takes years to develop, probably reflecting the ability of the parasites to evade immunity. A recent example of this is the binding of the Fc region of IgM to VAR2CSA‐type PfEMP1. This interferes with specific IgG recognition and phagocytosis of opsonized infected erythrocytes (IEs) without compromising the placental IE adhesion mediated by this PfEMP1 type. IgM also binds via Fc to several other PfEMP1 proteins, where it has been proposed to facilitate rosetting (binding of uninfected erythrocytes to a central IE). To further dissect the functional role of Fc‐mediated IgM binding to PfEMP1, we studied the PfEMP1 protein HB3VAR06, which mediates rosetting and binds IgM. Binding of IgM to this PfEMP1 involved the Fc domains Cμ3‐Cμ4 in IgM and the penultimate DBL domain (DBLζ2) at the C‐terminus of HB3VAR06. However, IgM binding did not inhibit specific IgG labelling of HB3VAR06 or shield IgG‐opsonized IEs from phagocytosis. Instead, IgM was required for rosetting, and each pentameric IgM molecule could bind two HB3VAR06 molecules. Together, our data indicate that the primary function of Fc‐mediated IgM binding in rosetting is not to shield IE from specific IgG recognition and phagocytosis as in VAR2CSA‐type PfEMP1. Rather, the function appears to be strengthening of IE–erythrocyte interactions. In conclusion, our study provides new evidence on the molecular details and functional significance of rosetting, a long‐recognized marker of parasites that cause severe P
. falciparum malaria.
A novel and generally applicable method for determining structures of membrane proteins in solution via small-angle neutron scattering (SANS) is presented. Common detergents for solubilizing membrane proteins were synthesized in isotope-substituted versions for utilizing the intrinsic neutron scattering length difference between hydrogen and deuterium. Individual hydrogen/deuterium levels of the detergent head and tail groups were achieved such that the formed micelles became effectively invisible in heavy water (D O) when investigated by neutrons. This way, only the signal from the membrane protein remained in the SANS data. We demonstrate that the method is not only generally applicable on five very different membrane proteins but also reveals subtle structural details about the sarco/endoplasmatic reticulum Ca ATPase (SERCA). In all, the synthesis of isotope-substituted detergents makes solution structure determination of membrane proteins by SANS and subsequent data analysis available to nonspecialists.
64Cu radiolabelled nanodiscs based on the 11 α-helix MSP1E3D1 protein and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine lipids were, for the first time, followed in vivo by positron emission tomography for evaluating the biodistribution of nanodiscs. A cancer tumor bearing mouse model was used for the investigations, and it was found that the approximately 13 nm nanodiscs, due to their size, permeate deeply into cancer tissue. This makes them promising candidates for both drug delivery purposes and as advanced imaging agents. For the radiolabelling, a simple approach for 64Cu radiolabelling of proteins via a chelating agent, DOTA, was developed. The reaction was performed at sufficiently mild conditions to be compatible with labelling of the protein part of a lipid-protein particle while fully conserving the particle structure including the amphipathic protein fold.
Targeted delivery of chemotherapeutics to cancer cells has the potential to yield high drug concentrations in cancer cells while minimizing any unwanted side effects. However, the development of multidrug resistance...
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