Comparative Study of Active and Allosteric Interaction in Protein Kinases

2021 ◽  
Vol 8 (1) ◽  
pp. 23-31
Author(s):  
Jefrin Ahmed ◽  
Judith Mary Lamo ◽  
Baphilinia Jones Mylliemngap
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Kinases ◽  
Active Site ◽  
Cell Function ◽  
Binding Capacity ◽  
Second Messengers ◽  
Gene Families ◽  
Allosteric Site ◽  
Kinase A

Protein kinases are key regulators of cell function that constitute one of the largest and most functionally diverse gene families. By adding phosphate groups to substrate proteins, they direct the activity, localization and overall function of many proteins, and serve to orchestrate the activity of almost all cellular processes. The main protein kinases consist of protein kinase A (PKA), protein kinase B (PKB), and protein kinase C (PKC) and are distinguished from each other by the different intracellular second messengers involved in their regulation and by the selective substrates they use. They all have a binding site for Mg2+-ATP (phosphate donor) and for substrate protein as well as various regulatory sites. We formulated to compare the binding capacity of protein kinases at the active site to allosteric sites. By comparing the active site and allosteric site of the protein kinases – A, B and C, using molecular docking it was found that in most of the cases the binding energy is high when an inhibitor binds to an active site as compared to the allosteric site. This comparison gave us an understanding of the interaction and inhibition of compounds to protein kinases in order to inhibit the activity of protein kinase A, B and C. It was concluded that for inhibiting the protein kinase function such as cell division and proliferation, binding of inhibitor to the allosteric site will be more effective.

A-kinase-anchoring proteins and cytoskeletal signalling events

2003 ◽  
Vol 31 (1) ◽  
pp. 87-89 ◽  
Author(s):  
J.D. Scott
Keyword(s):  
Protein Kinase ◽  
Molecular Mass ◽  
Protein Kinase A ◽  
Protein Kinases ◽  
Recent Progress ◽  
Membrane Receptors ◽  
Anchoring Proteins ◽  
Syndrome Protein ◽  
Kinase A

Targeting of protein kinases and phosphatases to the cytoskeleton enhances the regulation of many signalling events. Cytoskeletal signalling complexes facilitate this process by optimizing the relay of messages from membrane receptors to specific sites on the actin cytoskeleton. These signals influence fundamental cell properties such as shape, movement and division. Targeting of the cAMP-dependent kinase (protein kinase A) and other enzymes to this compartment is achieved through interaction with A-kinase-anchoring proteins (AKAPs). The present paper discusses recent progress on dissecting the biological role of WAVE1 (Wiskott–Alrich syndrome protein family verprolin homology protein 1), an AKAP that assembles a cytoskeletal transduction complex in response to signals that emanate from the low-molecular-mass GTPase, Rac.

scholarly journals Potential Role for Protein Kinases in Regulation of Bidirectional Endoplasmic Reticulum-to-Golgi Transport Revealed by Protein Kinase Inhibitor H89

2000 ◽  
Vol 11 (8) ◽  
pp. 2577-2590 ◽  
Author(s):  
Tina H. Lee ◽  
Adam D. Linstedt
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Kinases ◽  
Potential Role ◽  
Kinase Inhibitor ◽  
Transport Processes ◽  
Protein Kinase Inhibitors ◽  
Er Export ◽  
Kinase A

Recent evidence suggests a regulatory connection between cell volume, endoplasmic reticulum (ER) export, and stimulated Golgi-to-ER transport. To investigate the potential role of protein kinases we tested a panel of protein kinase inhibitors for their effect on these steps. One inhibitor, H89, an isoquinolinesulfonamide that is commonly used as a selective protein kinase A inhibitor, blocked both ER export and hypo-osmotic-, brefeldin A-, or nocodazole-induced Golgi-to-ER transport. In contrast, H89 did not block the constitutive ER Golgi-intermediate compartment (ERGIC)-to-ER and Golgi-to-ER traffic that underlies redistribution of ERGIC and Golgi proteins into the ER after ER export arrest. Surprisingly, other protein kinase A inhibitors, KT5720 and H8, as well as a set of protein kinase C inhibitors, had no effect on these transport processes. To test whether H89 might act at the level of either the coatomer protein (COP)I or the COPII coat protein complex we examined the localization of βCOP and Sec13 in H89-treated cells. H89 treatment led to a rapid loss of Sec13-labeled ER export sites but βCOP localization to the Golgi was unaffected. To further investigate the effect of H89 on COPII we developed a COPII recruitment assay with permeabilized cells and found that H89 potently inhibited binding of exogenous Sec13 to ER export sites. This block occurred in the presence of guanosine-5′-O-(3-thio)triphosphate, suggesting that Sec13 recruitment is inhibited at a step independent of the activation of the GTPase Sar1. These results identify a requirement for an H89-sensitive factor(s), potentially a novel protein kinase, in recruitment of COPII to ER export sites, as well as in stimulated but not constitutive Golgi-to-ER transport.

scholarly journals Discovery of cellular substrates for protein kinase A using a peptide array screening protocol

2011 ◽  
Vol 438 (1) ◽  
pp. 103-110 ◽  
Author(s):  
F. Donelson Smith ◽  
Bret K. Samelson ◽  
John D. Scott
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Kinases ◽  
Covalent Modification ◽  
Peptide Arrays ◽  
Post Translational Modification ◽  
Cellular Events ◽  
Screening Protocol ◽  
Full Complement ◽  
Kinase A

Post-translational modification of proteins is a universal form of cellular regulation. Phosphorylation on serine, threonine, tyrosine or histidine residues by protein kinases is the most widespread and versatile form of covalent modification. Resultant changes in activity, localization or stability of phosphoproteins drives cellular events. MS and bioinformatic analyses estimate that ~30% of intracellular proteins are phosphorylated at any given time. Multiple approaches have been developed to systematically define targets of protein kinases; however, it is likely that we have yet to catalogue the full complement of the phosphoproteome. The amino acids that surround a phosphoacceptor site are substrate determinants for protein kinases. For example, basophilic enzymes such as PKA (protein kinase A), protein kinase C and calmodulin-dependent kinases recognize basic side chains preceding the target serine or threonine residues. In the present paper we describe a strategy using peptide arrays and motif-specific antibodies to identify and characterize previously unrecognized substrate sequences for protein kinase A. We found that the protein kinases PKD (protein kinase D) and MARK3 [MAP (microtubule-associated protein)-regulating kinase 3] can both be phosphorylated by PKA. Furthermore, we show that the adapter protein RIL [a product of PDLIM4 (PDZ and LIM domain protein 4)] is a PKA substrate that is phosphorylated on Ser119 inside cells and that this mode of regulation may control its ability to affect cell growth.

scholarly journals Phosphorylation-Dependent SERS Readout for Activity Assay of Protein Kinase A in Cell Extracts

Nanomaterials ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 575
Author(s):  
Renyong Liu ◽  
Chenggen Xie ◽  
Yehan Yan ◽  
Lin Hu ◽  
Suhua Wang ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Biological Samples ◽  
Cell Function ◽  
Activity Assay ◽  
Cell Extracts ◽  
Disease Diagnostics ◽  
Wide Range ◽  
Surface Enhanced Raman ◽  
Kinase A

Protein kinases are key regulators of cell function, the abnormal activity of which may induce several human diseases, including cancers. Therefore, it is of great significance to develop a sensitive and reliable method for assaying protein kinase activities in real biological samples. Here, we report the phosphorylation-dependent surface-enhanced Raman scattering (SERS) readout of spermine-functionalized silver nanoparticles (AgNPs) for protein kinase A (PKA) activity assay in cell extracts. In this assay, the presence of PKA would phosphorylate and alter the net charge states of Raman dye-labeled substrate peptides, and the resulting anionic products could absorb onto the AgNPs with cationic surface charge through electrostatic attraction. Meanwhile, the Raman signals of dyes labeled on peptides were strongly enhanced by the aggregated AgNPs with interparticle hot spots formed in assay buffer. The SERS readout was directly proportional to the PKA activity in a wide range of 0.0001–0.5 U·μL−1 with a detection limit as low as 0.00003 U·μL−1. Moreover, the proposed SERS-based assay for the PKA activity was successfully applied to monitoring the activity and inhibition of PKA in real biological samples, particularly in cell extracts, which would be beneficial for kinase-related disease diagnostics and inhibitor screening.

Identification of a pathway by which glucose regulates β-catenin signalling via the cAMP/protein kinase A pathway in β-cell models

2013 ◽  
Vol 449 (3) ◽  
pp. 803-811 ◽  
Author(s):  
Emmanuelle Cognard ◽  
Coralie G. Dargaville ◽  
Deborah L. Hay ◽  
Peter R. Shepherd
Keyword(s):  
Insulin Secretion ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Cell Function ◽  
Regulation Of Gene Expression ◽  
Cell Models ◽  
Β Cells ◽  
Β Cell ◽  
Glucose Levels ◽  
Kinase A

Pancreatic β-cells are highly responsive to changes in glucose, but the mechanisms involved are only partially understood. There is increasing evidence that the β-catenin signalling pathway plays an important role in regulating β-cell function, but the mechanisms regulating β-catenin signalling in these cells is not well understood. In the present study we show that β-catenin levels and downstream signalling are regulated by changes in glucose levels in INS-1E and β-TC6-F7 β-cell models. We found a glucose-dependent increase in levels of β-catenin in the cytoplasm and nucleus of INS-1E cells. Expression of cyclin D1 also increased with glucose and required the presence of β-catenin. This was associated with an increase in phosphorylation of β-catenin on Ser552, which is known to stabilize the molecule and increase its transcriptional activity. In a search for possible signalling intermediates we found forskolin and cell-permeable cAMP analogues recapitulated the glucose effects, suggesting a role for cAMP and PKA (cAMP-dependent protein kinase/protein kinase A) downstream of glucose. Furthermore, glucose caused sustained increases in cAMP. Two different inhibitors of adenylate cyclase and PKA signalling blocked the effects of glucose, whereas siRNA (small interfering RNA) knockdown of PKA blocked the effects of glucose on β-catenin signalling. Finally, reducing β-catenin levels with either siRNA or pyrvinium impaired glucose- and KCl-stimulated insulin secretion. Taken together the results of the present study define a pathway by which changes in glucose levels can regulate β-catenin using a mechanism which involves cAMP production and the activation of PKA. This identifies a pathway that may be important in glucose-dependent regulation of gene expression and insulin secretion in β-cells.

Cross-talk between protein kinase A and the MAPK-activated protein kinases RSK1 and MK5

2010 ◽  
Vol 31 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Sergiy Kostenko ◽  
Alexey Shiryaev ◽  
Gianina Dumitriu ◽  
Nancy Gerits ◽  
Ugo Moens
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Kinases ◽  
Cross Talk ◽  
Kinase A
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