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D B McKay

Publications and source records attributed to D B McKay.

103 records · Page 6Linked to original sources
PubMed1982

Two helix DNA binding motif of CAP found in lac repressor and gal repressor.

Comparison of both the DNA and protein sequences of catabolite gene activator protein (CAP) with the sequences of lac and gal repressors shows significant homologies between a sequence that forms a two alpha-helix motif in CAP and sequences near the amino terminus of both repressors. This two-helix motif is thought to be involved in specific DNA sequence recognition by CAP. The region in lac repressor to which CAP is homologous contains many i-d mutations that are defective in DNA binding. Less significant sequence homologies between CAP and phage repressors and activators are also shown. The amino acid residues that are critical to the formation of the two-helix motif are conserved, while those residues expected to interact with DNA are variable. These observations suggest the lac and gal repressors also have a two alpha-helix structural motif which is involved in DNA binding and that this two helix motif may be generally found in many bacterial and phage repressors. We conclude that one major mechanism by which proteins can recognize specific base sequences in double stranded DNA is via the amino acid side chains of alpha-helices fitting into the major groove of B-DNA.

Amino Acid Sequence↗
PubMed1982

Structure of catabolite gene activator protein at 2.9-A resolution. Incorporation of amino acid sequence and interactions with cyclic AMP.

The amino acid sequence of the Escherichia coli catabolite gene activator protein has been fit into a 2.9-A resolution electron density map. Each subunit of the dimer consists of two structurally distinct domains. The larger NH2-terminal domain is seen to bind cyclic AMP and forms all of the contacts between the subunits. The cyclic AMP is completely buried between the interior of the "beta roll" structure of the large domain and a long alpha helix; it makes important hydrogen-bonding interactions with residues from both subunits. The guanidinium group of a buried Arg makes an internal salt link with the phosphate of cyclic AMP. The 6-amino group of adenine interacts simultaneously with both subunits. This interaction with both subunits and the fact that cyclic GMP and cyclic IMP do not activate catabolite gene activator protein suggest that the binding of cyclic AMP may alter the relative orientation of the two subunits, which in turn would change the structure of a DNA binding site that is presumed to span the two smaller domains. The distribution and nature of side chains in the small domain do not rule out the possibility that catabolite gene activator protein binds to left-handed B-DNA.

Allosteric Regulation↗
PubMed1982

X-ray grade crystals of diphtheria toxin.

Diphtheria toxin, complexed with the endogenous dinucleotide ApUp, has been crystallized under novel conditions involving high concentrations of both polyethylene glycol and salt. Among four crystal forms obtained, one was found suitable for high resolution structural analysis by x-ray diffraction. This form is triclinic (space group P1) with 2 molecules per unit cell; unit cell parameters are a = 70.8 A, b = 70.7 A, c = 65.3 A, alpha = 95.2 degrees, beta = 91.3 degrees, gamma = 99.7 degrees.

Crystallization↗
PubMed1982

Biochemical and biophysical characterization of insulin granules isolated from rat pancreatic islets by an iso-osmotic gradient.

The size and polydispersity of insulin granules isolated from rat pancreatic islets by centrifugation on a linear iso-osmotic gradient (300 mosM) have been characterized by quasi-elastic light scattering and photon correlation spectroscopy. The separation of granules by the linear gradient technique was compared directly to isolation on discontinuous gradients of hypertonic sucrose (300-1950 mosM) and the greater efficiency of separation assessed by parallel measurements of protein, insulin, cytochrome oxidase and beta-glucuronidase. Granules isolated from pancreatic beta-cells had a mean particle diameter of 342 nm, buoyant density of 1.104, hydrated mass of 23 femtograms and maximal insulin content of 8-9.6 . 10(5) molecules per granule.

Animals↗
PubMed1982

Structural similarity in the DNA-binding domains of catabolite gene activator and cro repressor proteins.

It is shown that there is a structural similarity between the presumed DNA-binding regions of the Escherichia coli catabolite gene activator protein ("CAP") and the cro repressor protein ("cro") from bacteriophage lambda. The correspondence between the two proteins is particularly striking for a structural unit consisting of two consecutive alpha-helices. The 24 alpha-carbon atoms that constitute the two-helical structural units in the two proteins can be superimposed with a root-mean-square disagreement of 1.1 A. It is shown that this agreement is very unlikely to be due to a chance correspondence. For both CAP activator and cro repressor proteins it is the second alpha-helix of the two-helical unit that has been proposed to bind within the major groove of left-handed or right-handed B DNA, respectively [McKay, D. B. & Steitz, T. A. (1981) Nature (London) 290, 744-749; Anderson, W. F., Ohlendorf, D. H., Takeda, Y. & Matthews, B. W. (1981) Nature (London) 290, 754-758]. The structural correspondence between CAP and cro seen here, together with other recent evidence of sequence homologies between cro, CAP, and other proteins that bind double-stranded DNA, suggests that the two-helical unit is likely to be a common feature of many DNA-binding proteins. The results also suggest that some principles of specific protein-double-stranded DNA interaction may be general and include recognition via alpha-helices fitting into the major groove of the DNA.

Bacteriophage lambda↗
PubMed1981

Structure of catabolite gene activator protein at 2.9 A resolution suggests binding to left-handed B-DNA.

The 2.9 A resolution crystal structure of Escherichia coli catabolite gene activator protein (CAP) complexed with cyclic AMP reveals two distinct structural domains separated by a cleft. The smaller carboxy-terminal domain is presumed to bind DNA while the amino-terminal domain is seen to bind cyclic AMP. Model building studies suggest that CAP binds to left-handed B-type DNA, contracting its major groove via two alpha-helices. It is possible that the CAP conversion of right- to left-handed DNA in a closed supercoil, is what activates transcription by RNA polymerase.

Carrier Proteins↗
PubMed1979

Substrate binding closes the cleft between the domains of yeast phosphoglycerate kinase.

Using small angle x-ray scattering from solutions of yeast phosphoglycerate kinase, we have measured the radius of gyration of the enzyme both in the presence and in the abscence of ligands. We find that the radius of gyration decreases by 1.09 +/- 0.34 A upon binding both substrates MgATP and 3-phosphoglycerate to form the ternary complex. Smaller decreases, at the limit of the precision of the measurement, were found for the separate binding of MgATP (0.30 +/- 0.50 A). Using computer modeling, it has been estimated that a substrate-induced cleft closure in phosphoglycerate kinase resulting from one lobe rotating 8-14 degrees relative to the other lobe lobe is consistent with this observed change in radius of gyration. We suggest, therefore, that the conformational change that results in the smaller radius of gyration for the ternary complex is a hinge motion of the two lobes which produces a closing of the cleft between the two lobes. The apparent similarity of the ligand-induced change in phosphoglycerate kinase to the cleft closure in hexokinase suggests that this kind of conformational change may prove to be a rather general kinase phenomenon (Bennett, W.S., and Steitz T.A. (1978) Proc. Natl. Acad. Sci. U.S.A. 75, 4848-4852; Anderson, C.M., Zucker, F.H., and Steitz, T.A. (1979) Science 204, 375-380).

Adenosine Triphosphate↗