Tracing the substrate translocation mechanism in P-glycoprotein.

P-glycoprotein (Pgp) is a prototypical ATP-binding cassette (ABC) transporter of great biological and clinical significance.Pgp confers cancer multidrug resistance and mediates the bioavailability and pharmacokinetics of many drugs (Juliano and Ling, 1976; Ueda et al., 1986; Sharom, 2011). Decades of structural and biochemical studies have provided insights into how Pgp binds diverse compounds (Loo and Clarke, 2000; Loo et al., 2009; Aller et al., 2009; Alam et al., 2019; Nosol et al., 2020; Chufan et al., 2015), but how they are translocated through the membrane has remained elusive. Here, we covalently attached a cyclic substrate to discrete sites of Pgp and determined multiple complex structures in inward- and outward-facing states by cryoEM. In conjunction with molecular dynamics simulations, our structures trace the substrate passage across the membrane and identify conformational changes in transmembrane helix 1 (TM1) as regulators of substrate transport. In mid-transport conformations, TM1 breaks at glycine 72. Mutation of this residue significantly impairs drug transport of Pgp in vivo, corroborating the importance of its regulatory role. Importantly, our data suggest that the cyclic substrate can exit Pgp without the requirement of a wide-open outward-facing conformation, diverting from the common efflux model for Pgp and other ABC exporters. The substrate transport mechanism of Pgp revealed here pinpoints critical targets for future drug discovery studies of this medically relevant system.

Gallstone Ileus With Cholecystoenteric Fistula in an Elderly Female: A Case Report.

Mechanical small-bowel obstruction can occur due to various reasons, including the impaction of a gallstone in the ileum after it has passed through a cholecystoenteric fistula. Gallstone ileus is an infrequent yet significant cause of this condition. This case report documents an instance of gallstone ileus, which accounts for less than 1% of patients with mechanical small bowel obstruction. We report a 75-year-old female patient who presented with colicky pain in both upper quadrants, hyporexia, and constipation that worsened during a period of nine days, which subsequently was accompanied by nausea and vomiting of bilious appearance in the next three days. Abdominal CT reported a dilated common bile duct (1.7 cm) with multiple stones inside measuring between 5 and 8 mm associated with pneumobilia of intrahepatic bile ducts and dilatation of small intestinal loops produced by a high-density image of approximately 2.5 cm. Laparoscopic exploration showed an obstructive mass measuring 15 cm from the ileocecal valve corresponding to a 2.54 x 2.35 cm gallstone, which was removed and enterorrhaphy was performed. The sine qua non condition for gallstone ileus to occur is the presence of a fistula between the gallbladder and the gastrointestinal tract. The treatment is mainly surgical and should be aimed primarily at the intestinal obstruction and secondarily at the cholecystoenteric fistula. This condition tends to have a high rate of complications and consequently long hospital stays. Making a timely diagnosis provides us with the tools for a surgical approach aimed at intestinal obstruction and subsequently in the management of the biliary fistula.