A bipartite mechanism for ERK2 recognition by its cognate regulators and substrates.
Mitogen-activated protein (MAP) kinases control gene expression in response to extracellular stimuli and exhibit exquisite specificity for their cognate regulators and substrates. We performed a structure-based mutational analysis of ERK2 to identify surface areas that are important for recognition of its interacting proteins. We show that binding and activation of MKP3 by ERK2 involve two distinct protein-protein interaction sites in ERK2. Thus, the common docking (CD) site composed of Glu-79, Tyr-126, Arg-133, Asp-160, Tyr-314, Asp-316, and Asp-319 are important for high affinity MKP3 binding but not essential for ERK2-induced MKP3 activation. MKP3 activation requires residues Tyr-111, Thr-116, Leu-119, Lys-149, Arg-189, Trp-190, Glu-218, Arg-223, Lys-229, and His-230 in the ERK2 substrate-binding region, located distal to the common docking site. Interestingly, many of the residues important for MKP3 recognition are also used for Elk1 binding and phosphorylation. In addition to the shared residues, there are also residues that are unique to each target recognition. There is evidence indicating that the CD site and the substrate-binding region defined here are also utilized for MEK1 recognition, and indeed, we demonstrate that the binding of MKP3, Elk1, and MEK1 to ERK2 is mutually exclusive. Taken together, our data suggest that the efficiency and fidelity of ERK2 signaling is achieved by a bipartite recognition process. In this model, one part of the ERK2-binding proteins (e.g. the kinase interaction motif sequence) docks to the CD site located on the back side of the ERK2 catalytic pocket for high affinity association, whereas the interaction of the substrate-binding region with another structural element (e.g. the FXFP motif in MKP3 and Elk1) may not only stabilize binding but also provide contacts crucial for modulating the activity and/or specificity of ERK2 target molecules.
Zhang
Jialin
J
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Zhou
Bo
B
Zheng
Chao-feng
CF
Zhang
Zhong-yin
ZY
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D004330:Drosophila; D054642:Dual Specificity Phosphatase 6; D004789:Enzyme Activation; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D048369:MAP Kinase Kinase 1; D020935:MAP Kinase Signaling System; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1
D004268:DNA-Binding Proteins; D010455:Peptides; D011518:Proto-Oncogene Proteins; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D005982:Glutathione Transferase; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases; D054642:Dual Specificity Phosphatase 6; D017027:Protein Tyrosine Phosphatases