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Biomedical subjects

T E Creighton

Publications and source records attributed to T E Creighton.

At least 109 records · Page 6Linked to original sources
PubMed1981

Circular dichroism, Raman spectroscopy, and gel filtration of trapped folding intermediates of ribonuclease.

The intermediates of ribonuclease with one to four disulfide bonds trapped during refolding of the reduced protein have been compared to the fully reduced and native forms of the protein by gel filtration, circular dichroism, and Raman spectroscopy. Correctly refolded ribonuclease is indistinguishable from native protein, while a three-disulfide intermediate has a compact conformation which is very similar, but not identical. In contrast, all other intermediates with one to four disulfide bonds are qualitatively similar to fully reduced ribonuclease by their circular dichroism and Raman spectra, although the disulfide cross-links affect the dimensions of the polypeptide chain. The apparent absence of stable partially ordered structures in the initial disulfide intermediates implies that during refolding there are relatively few constraints on formation on disulfide bonds, although their formation is probably not entirely random. The stable conformation appears during refolding only when the three or four disulfide bonds capable of stabilizing a native-like conformation of the entire polypeptide chain occur simultaneously.

Chromatography, Gel↗
PubMed1980

Ultraviolet difference spectroscopy of intermediates trapped in unfolding and refolding of bovine pancreatic trypsin inhibitor.

The solvent surface accessibilities of the aromatic amino acids of bovine pancreatic trypsin inhibitor have been examined after the protein has been trapped at various stages of unfolding and refolding. Two types of near-ultraviolet difference spectroscopy were used in making these measurements. One type compares the near-ultraviolet spectrum of each protein with the spectrum of the native inhibitor; the other is solvent perturbation spectroscopy. The two types of difference spectroscopy utilized are compared and found to be equivalent measures of tyrosine solvent exposure if the perturbation spectra are corrected for a probable contribution by buried residues. The experimental values for tyrosine solvent exposure in the native inhibitor are in agreement with those calculated from its crystal structure. The results of these studies identify an order in which the four tyrosines and the four phenylalanines of bovine pancreatic trypsin inhibitor are removed from solvent as refolding proceeds. The relative solvent accessibilities of the aromatic residues suggest an ordering in which the protein chain obtains compact globular structures.

Animals↗
PubMed1980

Multiple binding of antibodies to antigens: effect on radioimmunoassay binding curves.

Factors affecting the binding curves measured by radioimmunoassay are examined by comparing theoretical binding curves calculated for ideal populations of homogeneous antibodies and antigens in order to test the conclusions of Berzofsky et al. [Berzofsky, J. A., Curd, J. G., & Schechter, A. N. (1976) Biochemistry 15, 2113--2121] about the effect of multiple antigenic determinants on the shapes of such curves. When such ideal binding curves with the same midpoints and limiting values expected for univalent and multivalent antigens are compared, it is clear that multivalent antigens do not produce abnormally steep binding curves but rather tend somewhat to produce less steep curves. Under practical experimental circumstances, the binding curves for univalent and multivalent antigens are predicted to be indistinguishable in shape. Some alternative causes of abnormally steep binding curves are suggested.

Antigen-Antibody Complex↗
PubMed1980

A conformational isomer of bovine pancreatic trypsin inhibitor protein produced by refolding.

Refolding of the bovine pancreatic trypsin inhibitor protein (BPTI) after reduction of its three cysteine disulphide linkages can occur when the reduced protein is placed in a suitable oxidizing medium (ref. 1 and refs therein). The refolding process has been studied by trapping chemically intermediates which contain only one or two disulphide bonds (ref. 1 and refs therein). We report here that during structural studies of these intermediates by NMR, we have found that complete re-oxidation of the protein results in substantial quantities of a metastable folded species which is identical to native BPTI in its covalent bonding (including the disulphide bonds) but possesses a somewhat different conformation. The existence of such a species is supported by circular dichroism measurements on refolded BPTI. This novel form of BPTI is of considerable interest because it can be used to provide information about the folding mechanism and conformational stability of the protein.

Animals↗
PubMed1980

Counting integral numbers of amino acid residues per polypeptide chain.

Proteins have integral numbers of each of the 20 amino acids. However, all the currently accepted methods of determining this number measure the ratio of moles of amino acid residue per mole of protein. This value is rarely close to an integer, due to experimental errors in determination of the molar amounts of both amino acid residues and polypeptide chain. A simple method which gives integral values of amino acid residues per polypeptide chain, independent of any other properties of the protein, would be useful in characterising proteins. We describe here one method; it is illustrated for the case of Cys residues, although the approach should be useful for many of the other 19 usual amino acids.

Amino Acids↗