Cryo-EM structure of a eukaryotic zinc transporter at a low pH suggests its Zn2+-releasing mechanism.

Zinc transporter 8 (ZnT8) is mainly expressed in pancreatic islet ß cells and is responsible for H+-coupled uptake (antiport) of Zn2+ into the lumen of insulin secretory granules. Structures of human ZnT8 and its prokaryotic homolog YiiP have provided structural basis for constructing a plausible transport cycle for Zn2+. However, the mechanistic role that protons play in the transport process remains unclear. Here we present a lumen-facing cryo-EM structure of ZnT8 from Xenopus tropicalis (xtZnT8) in the presence of Zn2+ at a luminal pH (5.5). Compared to a Zn2+-bound xtZnT8 structure at a cytosolic pH (7.5), the low-pH structure displays an empty transmembrane Zn2+-binding site with a disrupted coordination geometry. Combined with a Zn2+-binding assay our data suggest that protons may disrupt Zn2+ coordination at the transmembrane Zn2+-binding site in the lumen-facing state, thus facilitating Zn2+ release from ZnT8 into the lumen.

Metal Versus Plastic Stents for Pancreatic Fluid Collection Drainage: A Systematic Review and Meta-analysis.

OBJECTIVES: The therapeutic efficacy of metal stents (MSs) for pancreatic fluid collections (PFCs) is invariably controversial. Here, we conducted a meta-analysis to summarize the results of efficacy of MSs and plastic stents (PSs) in PFC drainage. SUBJECTS AND METHODS: We performed a literature search of PubMed/MEDLINE, EMBASE, and COCHRANE for all of the published studies regarding the use of MSs and PSs for endoscopic transmural drainage of PFCs from January, 1 2015 to June 1, 2020. We extracted data from 9 studies (1359 patients) that met the inclusion criteria. The main outcome measures were the rates of treatment success, including technique success and clinical success (CS), adverse events, recurrence, procedure time, and length of hospital stay (LOS). RESULTS: There was no difference in overall technique success between patients treated with MSs and PSs for PFCs. However, MSs showed a higher CS rate 92% versus 82% (P<0.01) and a lower overall adverse event rate 20% versus 31% (P<0.01) than PSs. The recurrence rate of PFCs using MSs also had significant advantages over PSs 3% versus 10% (P<0.01) and MSs needed a shorter procedure time than PSs (26.73 vs. 45.40 min, P<0.01). In comparing direct endoscopic necrosectomy use and LOS, there was no difference between MSs and PSs. CONCLUSIONS: Bringing together the results of the current study, endoscopic ultrasound-guided drainage of PFCs using MSs may be superior to PSs in terms of CS, adverse events rates and recurrence rate, with similar LOS and direct endoscopic necrosectomy use.

Insight into binding of endogenous neurosteroid ligands to the sigma-1 receptor.

The sigma-1 receptor (σ1R) is a non-opioid membrane receptor, which responds to a diverse array of synthetic ligands to exert various pharmacological effects. Meanwhile, candidates for endogenous ligands of σ1R have also been identified. However, how endogenous ligands bind to σ1R remains unknown. Here, we present crystal structures of σ1R from Xenopus laevis (xlσ1R) bound to two endogenous neurosteroid ligands, progesterone (a putative antagonist) and dehydroepiandrosterone sulfate (DHEAS) (a putative agonist), at 2.15-3.09 Å resolutions. Both neurosteroids bind to a similar location in xlσ1R mainly through hydrophobic interactions, but surprisingly, with opposite binding orientations. DHEAS also forms hydrogen bonds with xlσ1R, whereas progesterone interacts indirectly with the receptor through water molecules near the binding site. Binding analyses are consistent with the xlσ1R-neurosteroid complex structures. Furthermore, molecular dynamics simulations and structural data reveal a potential water entry pathway. Our results provide insight into binding of two endogenous neurosteroid ligands to σ1R.

Engineering of a mammalian VMAT2 for cryo-EM analysis results in non-canonical protein folding.

Vesicular monoamine transporter 2 (VMAT2) belongs to the major facilitator superfamily (MFS), and mediates cytoplasmic monoamine packaging into presynaptic vesicles. Here, we present two cryo-EM structures of VMAT2, with a frog VMAT2 adopting a canonical MFS fold and an engineered sheep VMAT2 adopting a non-canonical fold. Both VMAT2 proteins mediate uptake of a selective fluorescent VMAT2 substrate into cells. Molecular docking, substrate binding and transport analysis reveal potential substrate binding mechanism in VMAT2. Meanwhile, caution is advised when interpreting engineered membrane protein structures.