Abstract. Hepatocyte Growth Factor (HGF, also known as Scatter Factor) is a powerful mitogen or motility factor in different cells, acting through the tyrosine kinase receptor encoded by the MET protooncogene. Endothelial cells express the MET gene and expose at the cell surface the mature protein (p190 MEt) made of a 50 kD (o0 subunit disulfide linked to a 145-kD (~) subunit. HGF binding to endothelial ceils identifies two sites with different affinities. The higher affinity binding site (Kd = 0.35 nM) corresponds to the p190 MET receptor. Sub-nanomolar concentrations of HGF, but not of a recombinant inactive precursor, stimulate the receptor kinase activity, cell proliferation and motility. HGF induces repairs of a wound in endothelial cell monolayer. HGF stimulates the scatter of endothelial cells grown on three-dimensional collagen gels, inducing an elongated phenotype. In the rabbit cornea, highly purified HGF promotes neovascularization at sub-nanomolar concentrations. HGF lacks activities related to hemostasis-thrombosis, inflammation and endothelial cells accessory functions. These data show that HGF is an in vivo potent angiogenic factor and in vitro induces endothelial cells to proliferate and migrate.
X.Zhang and S.Moréra contributed equally to this workThe BRCT domain (BRCA1 C-terminus), first identified in the breast cancer suppressor protein BRCA1, is an evolutionarily conserved protein-protein interaction region of~95 amino acids found in a large number of proteins involved in DNA repair, recombination and cell cycle control. Here we describe the first threedimensional structure and fold of a BRCT domain determined by X-ray crystallography at 3.2 Å resolution. The structure has been obtained from the C-terminal region of the human DNA repair protein XRCC1, and comprises a four-stranded parallel β-sheet surrounded by three α-helices, which form an autonomously folded domain. The compact XRCC1 structure explains the observed sequence homology between different BRCT motifs and provides a framework for modelling other BRCT domains. Furthermore, the established structure of an XRCC1 BRCT homodimer suggests potential protein-protein interaction sites for the complementary BRCT domain in DNA ligase III, since these two domains form a stable heterodimeric complex. Based on the XRCC1 BRCT structure, we have constructed a model for the C-terminal BRCT domain of BRCA1, which frequently is mutated in familial breast and ovarian cancer. The model allows insights into the effects of such mutations on the fold of the BRCT domain.
Conditions for the overexpression of human wild-type p53 using a baculovirus construct were optimised in insect cells which produced up to 20 mg p53/1 culture. Milligram amounts of p53 were purified to apparent homogeneity using chromatography on double-stranded DNA-cellulose (=58 % yield) followed by immunoaffinity chromatography with an epitope elution step (up to 48% yields) at 4°C. The M, of extracted p53 both from insect cell lysates and after purification was 54000 by SDSPAGE. Isoelectric focusing showed recombinant p53 to be an acidic protein, focusing at PI 6.0 under non-denaturing conditions. Expressed p53 at all stages of purification reacted by immunoblotting with specific p53 monoclonal antibodies, indicating the presence of intact epitopes at the Cterminus, N-terminus and central region of the protein. From ultracentrifugation studies, pure p53 exhibited significant oligomerisation, and sedimented broadly within the 7 -12-S region of sucrose gradients. Pure p53 slowly precipitated out of solution at concentrations between 1-6 mg/ml even in the presence of 1% detergent. Using metal affinity chromatography, we have established that pure p53 binds the immobilised divalent ions Zn2+, Ni2+ and Co2+ with high affinity.
Summary. Using a monoclonal antibody raised against an oestrogen receptor‐related protein, p29, and an indirect immunoperoxidase technique to stain human tissue, the presence of the antigen was investigated in normal, dysplastic and malignant tissue of the uterine cervix. In normal tissue p29 was present throughout the ectocervix during the menstrual cycle and virtually absent from the endocervix. In dysplastic cervical tissue there was a decreasing p29 content with increasing severity of the dysplasia, and very low levels were seen in the carcinomatous tissues.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.