Improvement of photoaffinity SPR imaging platform and determination of the binding site of p62/SQSTM1 to p38 MAP kinase.
p38 mitogen-activated protein kinase (MAPK) is a member of the serine/threonine kinases and is activated in response to stress stimuli, such as cytokines, ultraviolet irradiation, heat shock, and osmotic shock. We revealed in a previous report that p62/SQSTM1, known to participate in proteasomal or autophagosomal protein degradation and cytokine receptor signal transduction pathways, binds to p38 to regulate specifically. Herein, we describe the improvement of the photoaffinity-thiol linker of our SPR imaging platform, which enabled us to determine the binding site of p62 to p38. SPR imaging experiments using a new photoaffinity linker 2 to immobilize the peptides derived from p62 on gold substrate indicate that the domain comprising amino acids 164-190 of p62 binds to p38 directly. These SPR analysis data and empirical biologic data reveal that the binding site of p62 to p38 is the domain corresponding to 173-182.
Saito
Akiko
A
Antibiotics Laboratory, Chemical Biology Department, Advanced Research Institute, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
Kawai
Kayoko
K
Takayama
Hiroshi
H
Sudo
Tatsuhiko
T
Osada
Hiroyuki
H
D048868:Adaptor Proteins, Signal Transducing / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D003432:Cross-Linking Reagents / Q000737:chemistry; D015394:Molecular Structure; D019920:Photoaffinity Labels / Q000737:chemistry*; D013438:Sulfhydryl Compounds / Q000737:chemistry; D020349:Surface Plasmon Resonance / Q000379:methods*; D048051:p38 Mitogen-Activated Protein Kinases / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D003432:Cross-Linking Reagents; D019920:Photoaffinity Labels; D013438:Sulfhydryl Compounds; D048051:p38 Mitogen-Activated Protein Kinases