Regiospecificity of aminoglycoside phosphotransferase from Enterococci and Staphylococci (APH(3')-IIIa)
UMass Chan Affiliations
Department of Biochemistry and Molecular PharmacologyDocument Type
Journal ArticlePublication Date
1996-07-02Keywords
AminoglycosidesAnti-Bacterial Agents
Carbohydrate Sequence
Drug Resistance, Microbial
Enterococcus
Kanamycin Kinase
Magnetic Resonance Spectroscopy
Molecular Sequence Data
Phosphorylation
Phosphotransferases (Alcohol Group Acceptor)
Spectrometry, Mass, Secondary Ion
Staphylococcus
Substrate Specificity
Biochemistry
Enzymes and Coenzymes
Medicinal-Pharmaceutical Chemistry
Therapeutics
Metadata
Show full item recordAbstract
The broad-spectrum aminoglycoside phosphotransferase, APH(3')-IIIa, confers resistance to several aminoglycoside antibiotics in opportunistic pathogens of the genera Staphylococcus and Enterococcus. The profile of the drug resistance phenotype suggested that the enzyme would transfer a phosphate group from ATP to the 3'-hydroxyl of aminoglycosides. In addition, resistance to the 3'-deoxyaminoglycoside antibiotic, lividomycin A, suggested possible transfer to the 5"-hydroxyl of the ribose [Trieu-Cuot, P., and Courvalin, P. (1983) Gene 23, 331-341]. Using purified overexpressed enzyme, we have prepared and purified the products of APH(3')-IIIa-dependent phosphorylation of several of aminoglycoside antibiotics. Mass spectral analysis revealed that 4,6-disubstituted aminocyclitol antibiotics such as amikacin and kanamycin are monophosphorylated, while 4,5-disubstituted aminoglycosides such as butirosin A, ribostamycin, and neomycin B are both mono- and diphosphorylated by APH(3')-IIIa. Using a series of one- and two-dimensional 1H, 13C, and 31P NMR experiments, we have unambiguously assigned the regiospecificity of phosphoryl transfer to several antibiotics. The 4,6-disubstituted aminocyclitol antibiotics are exclusively phosphorylated at the 3'-OH hydroxyl, and the 4,5-disubstituted aminocyclitol antibiotics can be phosphorylated at both the 3'- and 5"-hydroxyls. The first phosphorylation can occur on either the 3'- or 5"-hydroxyl group of neomycin B or butirosin A. Initial phosphotransfer to the 3'-position predominates for butirosin while the 5"-OH is favored for neomycin. These results open the potential for the rational design of aminoglycoside kinase inhibitors based on functionalization of either the 6-aminohexose or the pentose rings of aminoglycoside antibiotics.Source
Biochemistry. 1996 Jul 2;35(26):8686-95. Link to article on publisher's siteDOI
10.1021/bi960389wPermanent Link to this Item
http://hdl.handle.net/20.500.14038/50094Notes
At the time of publication, Paul Thompson was not yet affiliated with UMass Medical School.
Related Resources
Link to Article in PubMedae974a485f413a2113503eed53cd6c53
10.1021/bi960389w