Structural basis for the active site inhibition mechanism of human kidney-type glutaminase (KGA).
Glutaminase is a metabolic enzyme responsible for glutaminolysis, a process harnessed by cancer cells to feed their accelerated growth and proliferation. Among the glutaminase isoforms, human kidney-type glutaminase (KGA) is often upregulated in cancer and is thus touted as an attractive drug target. Here we report the active site inhibition mechanism of KGA through the crystal structure of the catalytic domain of KGA (cKGA) in complex with 6-diazo-5-oxo-L-norleucine (DON), a substrate analogue of glutamine. DON covalently binds with the active site Ser286 and interacts with residues such as Tyr249, Asn335, Glu381, Asn388, Tyr414, Tyr466 and Val484. The nucleophilic attack of Ser286 sidechain on DON releases the diazo group (N2) from the inhibitor and results in the formation of an enzyme-inhibitor complex. Mutational studies confirmed the key role of these residues in the activity of KGA. This study will be important in the development of KGA active site inhibitors for therapeutic interventions.
Department of Biological Sciences, 14 Science Drive 4, National University of Singapore, Singapore 117543.
K
K
Thangavelu
Department of Biological Sciences, 14 Science Drive 4, National University of Singapore, Singapore 117543.
Qing Yun
QY
Chong
1] Department of Biological Sciences, 14 Science Drive 4, National University of Singapore, Singapore 117543 [2] Mechanobiology Institute Singapore, National University of Singapore, Singapore 117411.
Boon Chuan
BC
Low
Department of Biological Sciences, 14 Science Drive 4, National University of Singapore, Singapore 117543.
J
J
Sivaraman
D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D003980:Diazooxonorleucine / Q000378:metabolism*; D005972:Glutaminase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005973:Glutamine / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D013379:Substrate Specificity
D003980:Diazooxonorleucine; D005973:Glutamine; D005972:Glutaminase