KINETICS AND REGULATION OF ERYTHROCYTE K-CL COTRANSPORT

红细胞 K-CL 协同转运的动力学和调节

• 批准号: 2139988
• 负责人: PETER K LAUF
• 金额: $ 15.9万
• 依托单位: WRIGHT STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-09-01 至 1997-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2139988
• 关键词: acid base balance; adenosine triphosphate; atomic absorption spectrometry; bicarbonates; cell morphology; chemical kinetics; chlorine; electrophoresis; erythrocyte membrane; erythrocytes; genotype; ion transport; ionophores; magnesium; maleimides; mathematical model; membrane activity; membrane permeability; membrane transport proteins; nuclear magnetic resonance spectroscopy; nystatin; potassium; protein structure function; radiotracer; reticulocytes; sheep; thermodynamics; thiols

This project proposes a study of the kinetic, thermodynamic and regulatory properties of K-Cl cotransport of sheep red blood cells (RBCs), during the reticulocyte/mature RBC transition, and in ghosts. K- Cl cotransport, a ouabain-resistant (OR) and strictly Cl-dependent potassium (k) transporter, when activated by cell swelling or thiol group modification, may constitute a major fraction of the membrane's passive K permeability. Outwardly poised, K-Cl cotransport has been implicated in maturational RBC volume reduction of sheep and other RBCs, and pathologically in RBC dehydration of patients with certain hemoglobinopathies. In sheep, K-Cl cotransport occurs in all reticulocytes and in mature RBCs of the low K (LK) type but disappears in high K (HK) cells, The mechanism by which K-Cl cotransport is maintained moderately active in LK, however, inactivated in HK cells, is unknown. To understand the process of K-Cl cotransport involution in this model the following hypothesis will be tested: 1. The kinetic and thermodynamic characteristics of K-Cl cotransport, recently elucidated by us for the swelling activated system, are also common to hemoglobin-free ghosts, and to both reticulocyte precursor and mature RBCs. 2. Regulation involves membrane components of the transporter, as well as cytoplasmic factors. These general hypotheses will be tested as follows. The kinetic and thermodynamic properties of K-Cl cotransport will be compared by OR zero-trans K influxes and effluxes in reticulocytes and mature red cells of both LK and HK genotypes, and in resealed ghosts with those of the swelling induced K-Cl pathway. The regulation of K-Cl cotransport will be studied in LK cells with varied intracellular Mg, anions, and Ph, interventions known to cause activation or inactivation of the transporter. The presence of membrane-bound thiols crucial for K- Cl cotransport function will be tested by radio-labelling of selectively protected thiols and other groups, and electrophoretic verification of labelled membrane components, as well as by quantitative correlation of tracer incorporation with simultaneously measured K-Cl cotransprot activity. Understanding the process of transport changes in this model will elucidate why for example in human RBCs homozygous for hemoglobin S, K-Cl cotransport remains active and thus contributes to irreversible sickling.

本项目提出了一项动力学、热力学和热力学的研究