Polyproline-rod approach to isolating protein targets of bioactive small molecules: isolation of a new target of indomethacin.
Identification of protein targets of bioactive small molecules has been a technical hurdle of chemical genetics. Here we report a polyproline-rod approach to isolating protein targets of small molecules from cell lysates. The results indicate that insertion of a long, rigid polyproline helix between a small-molecule bait and a biotin tag boosts the capacity of affinity purification and thereby permits isolation of low-abundance or low-affinity proteins. In the course of the proof-of-concept experiments, we isolated glyoxalase 1 (GLO1) as a new target of indomethacin, a widely used antiinflammatory drug. Molecular biological experiments suggest that inhibition of GLO1 enzyme activity is related to the clinically recognized beneficial side effects of the indomethacin family of nonsteroidal antiinflammatory drugs.
Sato
Shin-Ichi
S
The Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
Kwon
Youngjoo
Y
Kamisuki
Shinji
S
Srivastava
Neeta
N
Mao
Qian
Q
Kawazoe
Yoshinori
Y
Uesugi
Motonari
M
D000595:Amino Acid Sequence; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000378:metabolism* / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry; D002478:Cells, Cultured; D006801:Humans; D007213:Indomethacin / Q000378:metabolism* / Q000494:pharmacology; D007791:Lactoylglutathione Lyase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology
D000894:Anti-Inflammatory Agents, Non-Steroidal; D000970:Antineoplastic Agents; D002457:Cell Extracts; D010455:Peptides; D011506:Proteins; D034741:RNA, Small Interfering; C011083:polyproline; D007791:Lactoylglutathione Lyase; D007213:Indomethacin