RESUMO
Despite the critical roles of excited states in protein functions, they remain intractable for most structural studies because of their notably low populations. Chemical shifts for "invisible" states in slow exchange with the ground state are intuitively observed using nuclear magnetic resonance (NMR) chemical exchange saturation transfer (CEST) experiments. Here, we present a CEST NMR spectroscopy study for the observation of protein pseudocontact shifts (PCSs) of excited states, which carry valuable angular and distance information about these states. We first validate this approach, dubbed PCS-CEST, in the slow-exchange system of Abp1p SH3-Ark1p labeled with lanthanide ions, where the PCSs of the minor states resemble those of the holo-form ground state as expected. We further demonstrate that pre-existing folding transitional conformations of an FF domain exhibit remarkably lower PCS values than the ground state, which suggests that the low-populated ensemble is unfolded or largely unfolded. A higher resolution of PCSs of the minor states is achieved using our 1D selective CEST experiments. Thus, PCS-CEST provides an exquisite structural probe into the minor but functionally essential excited states.
RESUMO
Using the technique of monoclonal antibody-immunogold-silver staining (M-IGSS), the mononuclear cells in peripheral blood (PB) and cerebrospinal fluid (CSF) were screened for immunoreactivity against JBE virus antigen (JBEV-Ag), interleukin-2 receptor (Tac), class II human leucocyte antigen (HLA-DR) and interferon-r (IFN-r) in 17 ELISA diagnosed JEB patients and 17 control cases with non-inflammatory and non-immunological diseases. It was found that JBEV-Ag+ cells were present in PB (16/17) and CSF (12/17) of the JBE group, but absent in both the control group and another 5 cases with mumps virus meningitis. The percentages of Tac+, HLA-DR+ and IFN-r+ cells in JBE patients were higher than those in controls.
