ABSTRACT
The cell maintains its intracellular pH in a narrow physiological range and disrupting the pH-homeostasis could cause dysfunctional metabolic states. Anion exchanger 2 (AE2) works at high cellular pH to catalyze the exchange between the intracellular HCO3- and extracellular Cl-, thereby maintaining the pH-homeostasis. Here, we determine the cryo-EM structures of human AE2 in five major operating states and one transitional hybrid state. Among those states, the AE2 shows the inward-facing, outward-facing, and intermediate conformations, as well as the substrate-binding pockets at two sides of the cell membrane. Furthermore, critical structural features were identified showing an interlock mechanism for interactions among the cytoplasmic N-terminal domain and the transmembrane domain and the self-inhibitory effect of the C-terminal loop. The structural and cell-based functional assay collectively demonstrate the dynamic process of the anion exchange across membranes and provide the structural basis for the pH-sensitive pH-rebalancing activity of AE2.
Subject(s)
Anion Transport Proteins , Antiporters , Humans , Chloride-Bicarbonate Antiporters , Hydrogen-Ion Concentration , Cell Membrane/metabolism , Homeostasis , Antiporters/metabolism , Anion Transport Proteins/metabolism , Chlorides/metabolismABSTRACT
As the major component of cell membranes, phosphatidylcholine (PC) is synthesized de novo in the Kennedy pathway and then undergoes extensive deacylation-reacylation remodeling via Lands' cycle. The re-acylation is catalyzed by lysophosphatidylcholine acyltransferase (LPCAT) and among the four LPCAT members in human, the LPCAT3 preferentially introduces polyunsaturated acyl onto the sn-2 position of lysophosphatidylcholine, thereby modulating the membrane fluidity and membrane protein functions therein. Combining the x-ray crystallography and the cryo-electron microscopy, we determined the structures of LPCAT3 in apo-, acyl donor-bound, and acyl receptor-bound states. A reaction chamber was revealed in the LPCAT3 structure where the lysophosphatidylcholine and arachidonoyl-CoA were positioned in two tunnels connected near to the catalytic center. A side pocket was found expanding the tunnel for the arachidonoyl CoA and holding the main body of arachidonoyl. The structural and functional analysis provides the basis for the re-acylation of lysophosphatidylcholine and the substrate preference during the reactions.
Subject(s)
1-Acylglycerophosphocholine O-Acyltransferase/chemistry , Phospholipids/chemistry , 1-Acylglycerophosphocholine O-Acyltransferase/metabolism , Acyl Coenzyme A/chemistry , Acyl Coenzyme A/metabolism , Acylation , Animals , Catalytic Domain , Chickens , Cryoelectron Microscopy , Crystallography, X-Ray , Lysophosphatidylcholines/chemistry , Lysophosphatidylcholines/metabolism , Models, Molecular , Phospholipids/metabolism , Protein Multimerization , Structure-Activity Relationship , Substrate SpecificityABSTRACT
Retroperitoneal ectopic pregnancy (REP) is a rare type of pregnancy with retroperitoneal implantation of the embryo. Herein, we report a case of a 29-year-old female with REP admitted in our department. Moreover, we review the literatures published previously reporting REP with the aim to improve the understanding of diagnosis and treatment of REP.
Subject(s)
Pregnancy, Ectopic , Adult , Female , Humans , Pregnancy , Pregnancy, Ectopic/diagnostic imaging , Retroperitoneal Space/diagnostic imagingABSTRACT
The skin contains three primary layers: epidermis, dermis, and hypodermis. Separation of epidermal components from dermis (dermal-epidermal separation) is an important basic investigation technique for pharmacology, toxicology, and biology. There are different systems of epidermal separation, including typical methods of chemical, enzyme, heat, etc. Each approach has advantages versus disadvantages, and thus the appropriate method should be chosen for a given research question. Here we described the method of enzyme separation.
Subject(s)
Cell Separation/methods , Dermis/cytology , Endopeptidases/metabolism , Epidermal Cells/cytology , Trypsin/metabolism , Bacterial Proteins/metabolism , Humans , Paenibacillus polymyxa/enzymology , Skin/cytologyABSTRACT
The skin contains three primary layers: epidermis, dermis, and hypodermis. Separation of epidermal components from dermis (dermal-epidermal separation) is an important basic investigation technique for pharmacology, toxicology, and biology. There are different systems of epidermal separation, including typical methods of chemical, enzyme, heat, etc. Each approach has advantages versus disadvantages, and thus the appropriate method should be chosen for a given research question. Here we described the method of chemical separation.
Subject(s)
Cell Separation/methods , Dermis/cytology , Epidermal Cells/cytology , Thiocyanates/pharmacology , Humans , Skin/cytologyABSTRACT
The skin contains three primary layers: epidermis, dermis, and hypodermis. Separation of epidermal components from the dermis (dermal-epidermal separation) is an important basic investigation technique for pharmacology, toxicology, and biology. There are different systems of epidermal separation, including typical methods of chemical, enzyme, heat, etc. Each approach has advantages versus disadvantages, and thus the appropriate method should be chosen for a given research question. Here we described the method of heat separation.
