6FUX

Structure of aminoglycoside phosphotransferase APH(3'')-Id from Streptomyces rimosus ATCC10970 in complex with ADP and streptomycin

Summary for 6FUX

• Entry DOI: 10.2210/pdb6fux/pdb
• Related: 6FUC
• Descriptor: Aminoglycoside phosphotransferase, GLYCEROL, ADENOSINE-5'-DIPHOSPHATE, ... (5 entities in total)
• Functional Keywords: streptomyces rimosus, aminoglycoside phosphotransferase, antibiotic resistance, streptomycin, phosphorylation, atp binding, kinase activity, transferase
• Biological source: Streptomyces rimosus subsp. rimosus ATCC 10970
• Total number of polymer chains: 1
• Total formula weight: 30944.30

Authors

Boyko, K.M.,Nikolaeva, A.Y.,Korzhenevskiy, D.A.,Alekseeva, M.G.,Mavletova, D.A.,Zakharevich, N.V.,Rudakova, N.N.,Danilenko, V.N.,Popov, V.O. (deposition date: 2018-02-28, release date: 2019-03-20, Last modification date: 2024-01-17)

Primary citation

Alekseeva, M.G.,Boyko, K.M.,Nikolaeva, A.Y.,Mavletova, D.A.,Rudakova, N.N.,Zakharevich, N.V.,Korzhenevskiy, D.A.,Ziganshin, R.H.,Popov, V.O.,Danilenko, V.N.
Identification, functional and structural characterization of novel aminoglycoside phosphotransferase APH(3′′)-Id from Streptomyces rimosus subsp. rimosus ATCC 10970.
Arch.Biochem.Biophys., 671:111-122, 2019

PubMed Abstract: In this study, we identified a new gene (aph(3″)-Id) coding for a streptomycin phosphotransferase by using phylogenetic comparative analysis of the genome of the oxytetracycline-producing strain Streptomyces rimosus ATCC 10970. Cloning the aph(3″)-Id gene in E.coli and inducing its expression led to an increase in the minimum inhibitory concentration of the recombinant E.coli strain to streptomycin reaching 350 μg/ml. To evaluate the phosphotransferase activity of the recombinant protein APH(3″)-Id we carried out thin-layer chromatography of the putative P-labeled streptomycin phosphate. We also performed a spectrophotometric analysis to determine the production of ADP coupled to NADH oxidation. Here are the kinetic parameters of the streptomycin phosphotransferase APH(3″)-Id: K 80.4 μM, V 6.45 μmol/min/mg and k 1.73 s. We demonstrated for the first time the ability of the aminoglycoside phototransferase (APH(3″)-Id) to undergo autophosphorylation in vitro. The 3D structures of APH(3″)-Id in its unliganded state and in ternary complex with streptomycin and ADP were obtained. The structure of the ternary complex is the first example of this class of enzymes with bound streptomycin. Comparison of the obtained structures with those of other aminoglycoside phosphotransferases revealed peculiar structure of the substrate-binding pocket reflecting its specificity to a particular antibiotic.

PubMed: 31251922
DOI: 10.1016/j.abb.2019.06.008

Experimental method

X-RAY DIFFRACTION (1.65 Å)