项目来源

美国卫生和人类服务部基金(HHS)

项目主持人

DAHMS, NANCY M.

项目受资助机构

MEDICAL COLLEGE OF WISCONSIN

项目编号

5R01DK04266728

立项年度

2022

立项时间

未公开

研究期限

未知 / 未知

项目级别

国家级

受资助金额

609491.00美元

学科

BIOCHEMISTRY

学科代码

未公开

基金类别

Research Projects

Address ; Alpha-galactosidase ; Animal Model ; Binding ; Binding Sites ; Biochemical ; Biodistribution ; Biogenesis ; Biological Assay ; Catabolism ; Cell Death ; Cell Survival ; Cell physiology ; Cells ; Cellular Assay ; Child ; Complex ; Crystallization ; Data ; Data Reporting ; Differentiation and Growth ; Disease ; Dissociation ; Electron Microscopy ; Enzymes ; Excision ; FDA approved ; Fabry Disease ; Family Characteristics ; Functional disorder ; Genetic Diseases ; Glycoside Hydrolases ; Glycosphingolipids ; Goals ; Homeostasis ; Housing ; Human ; Hydrolase

参与者AI

张蒙;薛涵;刘建方;任罡

参与机构

未公开

项目标书摘要：Lysosomes perform degradative metabolism critical to many endocytic,phagocytic,and autophagicprocesses.Cation-independent mannose 6-phosphate receptor(CI-MPR)plays a vital role in thebiogenesis of lysosomes by delivering~60 different newly synthesized hydrolytic enzymes withmannose 6-phosphate(M6P)on their N-glycans to lysosomes.Lysosomal storage diseases(LSDs)are caused by mutations in lysosomal proteins,mainly enzymes,that result in defective catabolismand substrate accumulation.Characteristic of the family of~70 LSDs is their progressive anddebilitating nature due to their impact on multiple organ systems.Treatment is symptomatic for mostLSDs,with only 11 having FDA-approved therapies.CI-MPR's ability to internalize recombinant M6P-containing enzymes delivered to patients by bi-weekly intravenous infusion forms the basis of enzymereplacement therapy(ERT)for 9 of these therapies.However,structural knowledge of the interactionbetween CI-MPR and its cargo of~60 different lysosomal enzymes is lacking.CI-MPR also binds adiverse set of extracellular non-M6P-containing ligands that mediate CI-MPR's tumor suppressor roleand regulation of cell growth and differentiation.CI-MPR regulates plasma insulin-like growth factor 2(IGF2)levels,and binds three components of the plasminogen activation system,plasminogen,urokinase-type plasminogen activator receptor(uPAR),and tissue-type PA(tPA).However,limitedinformation is available concerning the molecular basis for CI-MPR's interaction with plasminogen anduPAR.Here we will expand upon our preliminary data that:1)revealed the crystal structure of CI-MPR's N-terminal region that houses several ligand binding sites,2)showed the first structural view ofCI-MPR's entire extracellular region by high-resolution electron microscopy(EM),and 3)identifiedconformational changes elicited by pH and ligand binding.The overall structure of CI-MPR's 2300-residue extracellular region comprised of 15 domains will be determined alone and bound to ligandsusing an integrated approach combining single particle EM,mass spectrometry,X-ray crystallographyand NMR spectroscopy.The first structure of a complex between CI-MPR and a lysosomal enzyme(Aim 1),plasminogen(Aim 2),and uPAR(Aim 2)will be elucidated.Kinetic analyses and cell-basedassays will evaluate allosteric effects of each of CI-MPR's ligands(Aim 2).The mechanism of acidicpH-dependent ligand dissociation,which is essential to the functioning of CI-MPR and other endocyticreceptors,will be probed using mutagenesis studies and NMR techniques(Aim 3).Our new rat modelof the LSD Fabry disease will be used to test novel lysosomal enzyme-IGF2 fusion constructs for theirability to reduce substrate accumulation(Aim 4).These studies will provide insight for the design ofimproved therapeutics for the treatment of LSDs,and novel inhibitors of plasminogen activation.

项目官员

SECHI,SALVATORE

项目持续时间

28 years

项目负责机构类型

SCHOOLS OF MEDICINE