Author Correction: Closing the loop with reactions at the N-terminus
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Structural basis for intrinsic strand displacement activity of mitochondrial DNA polymerase
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Active ice sheet conservation cannot stop the retreat of Sermeq Kujalleq glacier, Greenland
Active conservation of an ice sheet seeks to reduce ice sheet mass loss and sea level rise. Here we explore the response of Sermeq Kujalleq in Greenland to limiting warm water inflow to the fjord it terminates by raising the sill by an artificial barrier at its mouth. We asynchronously couple an ice sheet model with a fjord model, and simulate glacier evolution with varying climate scenarios from the year 2020 to 2100. The tallest barrier cools the fjord water and reduces melt at the ice front. But this has minor impacts on glacier retreat under SSP5-8.5 and SSP2-4.5. Cooling the atmospheric forcing to 1990s levels reduces glacier retreat, but even reducing water temperatures with a barrier cannot stabilize the glacier. The glacier seems to be in an unstoppable phase of marine ice sheet instability on a rapidly deepening retrograde sloping bed and in water much deeper than in 2000s.
Structural mechanisms of human sodium-coupled high-affinity choline transporter CHT1
Mammalian sodium-coupled high-affinity choline transporter CHT1 uptakes choline in cholinergic neurons for acetylcholine synthesis and plays a critical role in cholinergic neurotransmission. Here, we present the high-resolution cryo-EM structures of human CHT1 in apo, substrate- and ion-bound, hemicholinium-3-inhibited, and ML352-inhibited states. These structures represent three distinct conformational states, elucidating the structural basis of the CHT1-mediated choline uptake mechanism. Three ion-binding sites, two for Na+ and one for Cl–, are unambiguously defined in the structures, demonstrating that both ions are indispensable cofactors for high-affinity choline-binding and are likely transported together with the substrate in a 2:1:1 stoichiometry. The two inhibitor-bound CHT1 structures reveal two distinct inhibitory mechanisms and provide a potential structural platform for designing therapeutic drugs to manipulate cholinergic neuron activity. Combined with the functional analysis, this study provides a comprehensive view of the structural mechanisms underlying substrate specificity, substrate/ion co-transport, and drug inhibition of a physiologically important symporter.
Adeno-associated virus serotype 2 capsids with proteolytic cuts by trypsin remain intact and potent
Recombinant adeno-associated viral (AAV) vectors have emerged as prominent gene delivery vehicles for gene therapy. In the journey of an AAV vector, AAV vectors can be exposed to different proteolytic environments inside the production cells, during the cell lysis step, within the endosome, and finally inside the cell nucleus. The stability of a modified AAV serotype 2 (AAV2) capsid was evaluated via a proteolytic approach using trypsin and other proteases and both denaturing and non-denaturing analytical methods. Trypsin digestion of the AAV2 capsids resulted in clips of the capsid proteins at the C-terminus as confirmed by denaturing methods including SDS-PAGE, CE-SDS, Western blot, and RPLC-MS. It was found that the AAV2 capsid with clips not only remains structurally intact, as confirmed by non-denaturing methods including SEC, thermostability testing, and cryo-EM, but also remains potent, as confirmed in a cell-based potency assay. This finding reveals that AAV2 capsid with proteolytic cuts remains intact and potent since the icosahedral three-dimensional structural arrangement of AAV capsid proteins can protect the clipped fragment from being released from the capsid, such that the AAV capsid remains intact allowing for the functionality to be maintained to deliver the DNA in the host cell. Evaluation of AAV stability using a proteolytic approach and multiple denaturing and non-denaturing analytical methods can provide valuable information for engineering AAV capsids to develop AAV-based gene therapy.
T-cell receptor structures and predictive models reveal comparable alpha and beta chain structural diversity despite differing genetic complexity
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