title: Crystal structures of mouse autocrine motility factor in complex with carbohydrate phosphate inhibitors provide insight into structure-activity relationship of the inhibitors.
abstract: Autocrine motility factor (AMF), a tumor-secreted cytokine, stimulates cell migration in vitro and metastasis in vivo. AMF is identical to the extracellular cytokines neuroleukin and maturation factor and, interestingly, to the intracellular enzyme phosphoglucose isomerase. The cytokine activity of AMF is inhibited by carbohydrate phosphate compounds as they compete for AMF binding with the carbohydrate moiety of the AMF receptor (AMFR), which is a glycosylated seven transmembrane helix protein. Here, we report the first comprehensive high-resolution crystal structure analyses of the inhibitor-free form and the eight types of inhibitor (phosphate, erythrose 4-phosphate (E4P), arabinose 5-phosphate (A5P), sorbitol 6-phosphate (S6P), 6-phosphogluconic acid (6PGA), fructose 6-phosphate (F6P), glucose 6-phosphate (G6P), or mannose 6-phosphate (M6P)) complexes of mouse AMF (mAMF). We assayed the inhibitory activities of these inhibitors against the cytokine activity of mAMF. The inhibitory activities of the six-carbon sugars (G6P, F6P, M6P, and 6PGA) were found to be significantly higher than those of the four or five-carbon sugars (E4P or A5P). The inhibitory activities clearly depend on the length of the inhibitor molecules. A structural comparison revealed that a water-mediated hydrogen bond between one end of the inhibitor and a rigid portion of the protein surface in the shorter-chain inhibitor (E4P) complex is replaced by a direct hydrogen bond in the longer-chain inhibitor (6PGA) complex. Thus, to obtain a new compound with higher inhibitory activities against AMF, water molecules at the inhibitor binding site of AMF should be replaced by a functional group of inhibitors in order to introduce direct interactions with the protein surface. The present structure-activity relationship studies will be valuable not only for designing more effective AMF inhibitors but also for studying general protein-inhibitor interactions.
authors.affiliation: School of Pharmaceutical Sciences, Showa University, Shinagawa-ku, Tokyo, Japan. ntanaka@pharm.showa-u.ac.jp
authors.forename: Nobutada
authors.initials: N
authors.lastname: Tanaka
authors.forename: Arayo
authors.initials: A
authors.lastname: Haga
authors.forename: Noriko
authors.initials: N
authors.lastname: Naba
authors.forename: Katsura
authors.initials: K
authors.lastname: Shiraiwa
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Kusakabe
authors.forename: Kazunori
authors.initials: K
authors.lastname: Hashimoto
authors.forename: Tatsuyoshi
authors.initials: T
authors.lastname: Funasaka
authors.forename: Hisamitsu
authors.initials: H
authors.lastname: Nagase
authors.forename: Avraham
authors.initials: A
authors.lastname: Raz
authors.forename: Kazuo T
authors.initials: KT
authors.lastname: Nakamura
mesh_terms: D000818:Animals; D001665:Binding Sites; D002241:Carbohydrates / Q000737:chemistry*; D002465:Cell Movement; D018360:Crystallography, X-Ray; D005956:Glucose-6-Phosphate Isomerase / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010710:Phosphates / Q000737:chemistry*; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry; D013329:Structure-Activity Relationship
chemical_list: D002241:Carbohydrates; D010710:Phosphates; D021122:Protein Subunits; D005956:Glucose-6-Phosphate Isomerase