HKUST Library Institutional Repository Banner

HKUST Institutional Repository >
Chemistry >
CHEM Master Theses >

Please use this identifier to cite or link to this item:
Title: Macromolecular crowding effects on the activity of the extracellular signal regulated kinase 2
Authors: Cao, Yang
Issue Date: 2008
Abstract: Mitogen-activated protein kinases (MAP kinase) relay, amplify, and integrate extracellular signals with high fidelity and efficiency in a highly crowded intracellular environment where the total macromolecular concentration reaches 20-40%. However, in vitro investigations in dilute solution show that they have low specificity, phosphorylating both physiological and nonphysiological substrates without sufficient differentiation. This contradiction cannot be reconciled by the docking interaction between the kinase and their physiological substrates. In this study, we have used the comparative kinetic studies to probe and elucidate the effects of macromolecular crowding on the substrate recognition and catalytic efficiency of the MAP kinases, using extracellular signal regulated protein kinase 2 (ERK2) as a model. The steady-state kinetic parameters of ERK2 for Ets11-138 and myelin basic protein (MBP) were measured (in normal dilute buffer, kcat/Km(MBP) = 2.9 ± 0.4 μM-1 min-1, kcat/Km(Ets1) = 11.8 ± 0.3 μM-1 min-1; in 30% Ficoll 70 solution, kcat/Km(MBP) = 9.3 ± 1.3 μM-1 min-1, kcat/Km(Ets1) = 51.9 ± 8.1 μM-1 min-1). The tryptophan fluorescence studies of ERK2, MBP and Ets11-138 were performed. It was found that the macromolecular crowding agent significantly increases the catalytic efficiency (kcat/Km) of ERK2 to both physiological substrate, Ets11-138 and nonphysiological substrate, MBP, particularly at high concentrations from 20% to 30%, through inducing possibly structural changes in protein, but, as we expected, the differentiation of efficiency to these two substrates in 30% Ficoll 70 solution was enlarged from 4 to 5.6 folds.
Description: Thesis (M.Phil.)--Hong Kong University of Science and Technology, 2008
xiv, 60 leaves : ill. ; 30 cm
HKUST Call Number: Thesis CHEM 2008 Cao
Appears in Collections:CHEM Master Theses

Files in This Item:

File Description SizeFormat

All items in this Repository are protected by copyright, with all rights reserved.