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16 results on '"Brown, Nick R."'

3. Specificity determinants of recruitment peptides bound to phospho-CDK2/cyclin A

Author

Lowe, Edward D., Tews, Ivo, Cheng, Kin Yip, Brown, Nick R., Gul, Sheraz, Noble, Martin E. M., Gamblin, Steven J., and Johnson, Louise N.

Subjects
Biochemistry -- Research, Peptides -- Physiological aspects, Phosphorus compounds -- Physiological aspects, Cell cycle -- Physiological aspects, Protein kinases -- Physiological aspects, Biological sciences, Chemistry
Abstract

Research has been conducted on cyclin dependent protein kinase 2 in association with cyclin A. Results demonstrate that cyclin recruitment site accommodates hydrophobic residue and that this site contributes to recruitment peptide recognition.

Published
2002

5. The cyclin box fold: protein recognition in cell-cycle and transcription control

Author

Noble, Martin E.M., Endicott, Jane A., Brown, Nick R., and Johnson, Louise N.

Subjects
Cell cycle -- Analysis, Nucleotide sequence -- Analysis, Proteins -- Analysis, Genetic transcription -- Analysis, Biological sciences, Chemistry
Abstract

Regulation of both the cell cycle and gene transcription is essential for orderly progression of cell growth and division. Recent results on the structures of two cyclins, cyclin A and cyclin H, and two transcription factor mediator proteins, TFIIB and the A pocket region of the retinoblastoma tumour suppressor protein (Rb), show that they share domains with a strikingly similar [Alpha]-helical topology, despite remote sequence identity.

Published
1997

15. XenopusPhospho-CDK7/Cyclin H Expressed in Baculoviral-Infected Insect Cells

Author

Lawrie, Alison M., Tito, Paula, Hernandez, Helena, Brown, Nick R., Robinson, Carol V., Endicott, Jane A., Noble, Martin E.M., and Johnson, Louise N.

Abstract

The cyclin-dependent kinase-activating kinase (CAK) catalyzes the phosphorylation of the cyclin-dependent protein kinases (CDKs) on a threonine residue (Thr160 in human CDK2). The reaction is an obligatory step in the activation of the CDKs. In higher eukaryotes, the CAK complex has been characterized in two forms. The first consists of three subunits, namely CDK7, cyclin H, and an assembly factor called MAT1, while the second consists of phospho-CDK7 and cyclin H. Phosphorylation of CDK7 is essential for cyclin association and kinase activity in the absence of the assembly factor MAT1. The Xenopus laevisCDK7 phosphorylation sites are located on the activation segment of the kinase at residues Ser170 and at Thr176 (the latter residue corresponding to Thr160 in human CDK2). We report the expression and purification of X. laevisCDK7/cyclin H binary complex in insect cells through coinfection with the recombinant viruses, AcCDK7 and Accyclin H. Quantities suitable for crystallization trials have been obtained. The purified CDK7/cyclin H binary complex phosphorylated CDK2 and CDK2/cyclin A but did not phosphorylate histone H1 or peptide substrates based on the activation segments of CDK7 and CDK2. Analysis by mass spectrometry showed that coexpression of CDK7 with cyclin H in baculoviral-infected insect cells results in phosphorylation of residues Ser170 and Thr176 in CDK7. It is assumed that phosphorylation is promoted by kinase(s) in the insect cells that results in the correct, physiologically significant posttranslational modification. We discuss the occurrence of in vivophosphorylation of proteins expressed in baculoviral-infected insect cells.

Published
2001
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16. The Role of the Phospho-CDK2/Cyclin A Recruitment Site in Substrate Recognition.

Author

Kin-Yip Cheng, Noble, Martin E. M., Skamnaki, Vicky, Brown, Nick R., Lowe, Ed D., Kontogiannis, Luke, Shen, Kui, Cole, Philip A., Siligardi, Giuliano, and Johnson, Louise N.

Subjects
*CELL cycle, *CELL proliferation, *PROTEIN kinases, *METABOLISM, *GROWTH factors, *CHEMICAL affinity
Abstract

Phospho-CDK2/cyclin A, a kinase that is active in cell cycle S phase, contains an RXL substrate recognition site that is over 40 A from the catalytic site. The role of this recruitment site, which enhances substrate affinity and catalytic efficiency, has been investigated using peptides derived from the natural substrates, namely CDC6 and p107, and a bispeptide inhibitor in which the γ-phosphate of ATP is covalently attached by a linker to the CDC6 substrate peptide. X-ray studies with a 30-residue CDC6 peptide in complex with pCDK2/cyclin A showed binding of a dodecamer peptide at the recruitment site and a heptapeptide at the catalytic site, but no density for the linking 11 residues. Kinetic studies established that the CDC6 peptide had an 18-fold lower Km compared with heptapeptide substrate and that this effect required the recruitment peptide to be covalently linked to the substrate peptide. X-ray studies with the CDC6 bispeptide showed binding of the dodecamer at the recruitment site and the modified ATP in two alternative conformations at the catalytic site. The CDC6 bispeptide was a potent inhibitor competitive with both ATP and peptide substrate of pCDK2/cyclin A activity against a heptapeptide substrate (Ki = 0.83 n) but less effective against RXL-containing substrates. We discuss how localization at the recruitment site (KD 0.4 μ) leads to increased catalytic efficiency and the design of a potent inhibitor. The notion of a flexible linker between the sites, which must have more than a minimal number of residues, provides an explanation for recognition and discrimination against different substrates. [ABSTRACT FROM AUTHOR]

Published
2006
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