Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0271188 (Halo)
 461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The production of recombinant proteins such as the fibroblast growth factors (FGFs) is the key to establishing their function in cell communication. The production of recombinant FGFs in E. coli is limited, however, due to expression and solubility problems. HaloTag has been used as a fusion protein to introduce a genetically-encoded means for chemical conjugation of probes. We have expressed 11 FGF proteins with an N-terminal HaloTag, followed by a tobacco etch virus (TEV) protease cleavage site to allow release of the FGF protein. These were purified by heparin-affinity chromatography, and in some instances by further ion-exchange chromatography. It was found that HaloTag did not adversely affect the expression of FGF1 and FGF10, both of which expressed well as soluble proteins. The N-terminal HaloTag fusion was found to enhance the expression and yield of FGF2, FGF3 and FGF7. Moreover, whereas FGF6, FGF8, FGF16, FGF17, FGF20 and FGF22 were only expressed as insoluble proteins, their N-terminal HaloTag fusion counterparts (Halo-FGFs) were soluble, and could be successfully purified. However, cleavage of Halo-FGF6, -FGF8 and -FGF22 with TEV resulted in aggregation of the FGF protein. Measurement of phosphorylation of p42/44 mitogen-activated protein kinase and of cell growth demonstrated that the HaloTag fusion proteins were biologically active. Thus, HaloTag provides a means to enhance the expression of soluble recombinant proteins, in addition to providing a chemical genetics route for covalent tagging of proteins.
...
PMID:HaloTag is an effective expression and solubilisation fusion partner for a range of fibroblast growth factors. 2613 34

The range of biological outcomes generated by many signalling proteins in development and homeostasis is increased by their interactions with glycosaminoglycans, particularly heparan sulfate (HS). This interaction controls the localization and movement of these signalling proteins, but whether such control depends on the specificity of the interactions is not known. We used five fibroblast growth factors with an N-terminal HaloTag (Halo-FGFs) for fluorescent labelling, with well-characterized and distinct HS-binding properties, and measured their binding and diffusion in pericellular matrix of fixed rat mammary 27 fibroblasts. Halo-FGF1, Halo-FGF2 and Halo-FGF6 bound to HS, whereas Halo-FGF10 also interacted with chondroitin sulfate/dermatan sulfate, and FGF20 did not bind detectably. The distribution of bound FGFs in the pericellular matrix was not homogeneous, and for FGF10 exhibited striking clusters. Fluorescence recovery after photobleaching showed that FGF2 and FGF6 diffused faster, whereas FGF1 diffused more slowly, and FGF10 was immobile. The results demonstrate that the specificity of the interactions of proteins with glycosaminoglycans controls their binding and diffusion. Moreover, cells regulate the spatial distribution of different protein-binding sites in glycosaminoglycans independently of each other, implying that the extracellular matrix has long-range structure.
...
PMID:Selectivity in glycosaminoglycan binding dictates the distribution and diffusion of fibroblast growth factors in the pericellular matrix. 2700 90