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Aminoglycoside O-Phosphotransferases

Aminoglycoside O-Phosphotransferases (APH) catalyzes the ATP-dependent phosphorylation of key hydroxyl groups of aminoglycosides.  The APH family of kinases includes APH(2 ), APH(3 ), APH(3 ), APH(7 ), APH(4), APH(6), and APH(9).

The best-studied member of the family is APH(3 )-IIIa, which is generally found in gram-positive pathogens. The enzyme has been crystallized and the 3D structure has been determined bound with nucleotide and aminoglycoside substrates [1-3]. The monomeric subunit of this dimeric protein is structurally very similar to the serine/threonine/tyrosine protein and the inositide kinases that are common to eukaryotic signaling pathways such as murine cAMP dependent protein kinase and casein kinase.

The details of the molecular mechanism of phosphate transfer and substrate recognition have been well worked out []4-7], and this has resulted in efforts to identify small molecule inhibitors of the class. A second homolog, APH(3 )-IIa, has also been crystallized and resolved to 2.1-˚A resolution as well, and it shares extensive structural similarities with APH(3 )-IIIa [8].

Like the protein and inositide kinases, the 3D structures of APHs display two distinct domains: The N-terminal β-sheet region is responsible for ATP binding, and the α-helical C-terminal provides the aminoglycoside recognition site. The active site, where phosphate transfer occurs, lies at the interface of the two domains.

APHs can be inhibited by small molecules well known to inhibit protein and inositide kinases such as wortmanin [9-10].

 

 


1. Hon, W. C.; McKay, G. A.; Thompson, P. R.; Sweet, R. M.; Yang, D. S. C.; Wright, G. D.; Berghuis, A. M. Cell 1997, 89, 887.

2. Fong, D. H.; Berghuis, A. M. EMBO J 2002, 21, 2323.

3. Burk, D. L.; Hon, W. C.; Leung, A. K.; Berghuis, A. M. Biochemistry 2001, 40, 8756.

4. McKay, G. A.; Wright, G. D. Biochemistry 1996, 35.

5. Thompson, P. R.; Boehr, D. D.; Berghuis, A. M.; Wright, G. D. Biochemistry 2002, 41, 7001.

6. Boehr, D. D.; Farley, A. R.; Wright, G. D.; Cox, J. R. Chem. Biol. 2002, 9, 1209.

7. Boehr, D. D.; Farley, A. R.; LaRonde, F. J.; Murdock, T. R.; Wright, G. D.; Cox, J. R. Biochemistry 2005, 44, 12445.

8. Nurizzo, D.; Shewry, S. C.; Perlin, M. H.; Brown, S. A.; Dholakia, J. N.; Fuchs, R. L.; Deva, T.; Baker, E. N.; Smith, C. A. J. Mol. Biol. 2003, 327, 491.

9. Daigle, D. M.; McKay, G. A.; Wright, G. D. J. Biol. Chem. 1997, 272, 24755.

10. Boehr, D. D.; Lane, W. S.; Wright, G. D. Chem Biol 2001, 8, 791.

 

 

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