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The major proteins in the lumen of the endoplasmic reticulum (ER) are thought to function in Ca2+ sequestration or as "molecular chaperones" in the folding and assembly of membrane or secreted proteins. Based on the ability of many chaperones to bind selectively to unfolded proteins and to dissociate from them upon ATP hydrolysis, we developed an affinity(More)
Several amino groups of recombinant human erythropoietin are selectively cross-linked by specific cross-linkers including disuccinimidyl suberate or dithiobis(succinimidyl propionate). Intramolecular cross-linkings are obtained without significant change of the protein conformation using appropriate concentrations (0.2 mM) of the cross-linkers, which(More)
An extracellular domain of a human neurotrophin receptor trkB was expressed in Chinese hamster ovary cells and isolated as a glycoprotein possessing binding activity for brain-derived neurotrophic factor. The extracellular domain contains 398 amino acids and has a molecular weight of 60.6 kDa according to laser desorption mass spectrometry, indicating that(More)
We recently observed two 2,4-dinitrophenylhydrazine (DNPH)-reactive proteins of 40 and 120 kDa in the bronchoalveolar lavage fluids of rats exposed to >95% O(2) for 48 h. The N-terminal sequences of these proteins were both identical over 16 amino acids with rat beta-casein, which, in addition to its more common association with milk, is produced by(More)
The microheterogeneity of the carbohydrate structures on recombinant human erythropoietin (rHuEPO) expressed in Chinese hamster ovary cells has been evaluated by electrospray ionization (ESI) mass spectrometry (MS) of glycopeptide fragments. The microheterogeneity is largely associated with the presence or absence of terminal N-acetylneuraminic acid(More)
The extracellular domain of the human neurotrophin TRKB receptor expressed in Chinese hamster ovary cells is a highly glycosylated protein, possessing binding ability for brain-derived neurotrophic factor (BDNF). Two distinct ligand binding domains of TRKB were isolated from proteolytic digests of the receptor by affinity separation on immobilized BDNF. One(More)
Recombinant human brain-derived neurotrophic factor (BDNF) is now under extensive investigation because of its potential clinical applications. Radioactively labeled proteins are usually required to study receptor binding and pharmacokinetic properties of proteins. This study was undertaken to see if iodination affects the biological and conformational(More)
The prt gene, encoding a protease (Prt) from Streptomyces lividans TK24, was cloned and sequenced. An S. lividans host with plasmid-borne prt secreted 200 micrograms/ml of a 22-kDa Prt into the culture medium. Prt is classified as a metalloprotease since its activity is significantly inhibited by 1,10-phenanthroline or EDTA. The region upstream from prt(More)
Can one protein sequence encode two structures? Oxidative folding of human insulin-like growth factor 1 (IGF-1), a globular protein of 70 residues, is shown to yield two products of similar thermodynamic stability. This observation is of particular interest in light of the recent demonstration that two of the three disulfide bonds in native IGF-1 rearrange(More)
The leptin receptor (OB-R) is a member of the class I cytokine receptor family and mediates the weight regulatory effects of its ligand through interaction with cytoplasmic kinases. The extracellular domain of this receptor is comprised of two immunoglobulin-like and cytokine-receptor homology domains each and type III fibronectin domains. The extracellular(More)