RESUMEN
The first extracellular domain (ECD-1) of the corticotropin releasing factor (CRF) type 1 receptor, (CRFR1), is important for binding of CRF ligands. A soluble protein, mNT-CRFR1, produced by COS M6 cells transfected with a cDNA encoding amino acids 1--119 of human CRFR1 and modified to include epitope tags, binds a CRF antagonist, astressin, in a radioreceptor assay using [(125)I-d-Tyr(0)]astressin. N-terminal sequencing of mNT-CRFR1 showed the absence of the first 23 amino acids of human CRFR1. This result suggests that the CRFR1 protein is processed to cleave a putative signal peptide corresponding to amino acids 1--23. A cDNA encoding amino acids 24--119 followed by a FLAG tag, was expressed as a thioredoxin fusion protein in Escherichia coli. Following thrombin cleavage, the purified protein (bNT-CRFR1) binds astressin and the agonist urocortin with high affinity. Reduced, alkylated bNT-CRFR1 does not bind [(125)I-D-Tyr(0)]astressin. Mass spectrometric analysis of photoaffinity labeled bNT-CRFR1 yielded a 1:1 complex with ligand. Analysis of the disulfide arrangement of bNT-CRFR1 revealed bonds between Cys(30) and Cys(54), Cys(44) and Cys(87), and Cys(68) and Cys(102). This arrangement is similar to that of the ECD-1 of the parathyroid hormone receptor (PTHR), suggesting a conserved structural motif in the N-terminal domain of this family of receptors.
Asunto(s)
Hormona Liberadora de Corticotropina/genética , Secuencia de Aminoácidos , Animales , Células COS , Dicroismo Circular , Hormona Liberadora de Corticotropina/química , Hormona Liberadora de Corticotropina/aislamiento & purificación , ADN Complementario , Humanos , Datos de Secuencia Molecular , SolubilidadRESUMEN
A prospective, double-blind study of 392 patients randomized into four groups was performed to establish whether diagnostic intravenous urograms could be obtained with a lower dose of iodine when using the dimeric, non-ionic contrast medium iodixanol compared with the monomeric, non-ionic iohexol. Patients received iodixanol or iohexol containing either 9 or 12 g of iodine (gI). The primary parameter was the diagnostic quality of the 6 min film, assessed in a blinded fashion, by consensus, by four radiologists. Iodixanol at both doses was diagnostic in over 90% of cases. Iohexol was only diagnostic in 74% (9 gI) and 81.8% (12 gI). Pairwise comparisons revealed that iodixanol 9 gI was significantly better than both iohexol 9 gI (p = 0.0005) and 12 gI (p = 0.014). No significant difference was present for different doses within the same contrast medium group. Iodixanol resulted in poorer bladder distension than iohexol. Iodixanol caused significantly less discomfort than iohexol.