73-kDa Molecular Chaperone HSP73 Is a Direct Target of Antibiotic Gentamicin

Abstract

Although gentamicin (GM) has been used widely as an antibiotic, the specific binding protein of the drug has not yet been understood sufficiently. Here we show that GM specifically associates with the 73-kDa molecular chaperone HSP73 and reduces its chaperone activity in vitro. In the present study, we investigated GM-specific binding proteins using a GM-affinity column and porcine kidney cytosol. After washing the column, only the 73-kDa protein was eluted from the column by the addition of 10 mM GM. None of the other proteins were found in the eluant. Upon immunoblotting, the protein was identical to HSP73. Upon CD spectrum analysis, the binding of GM to HSP73 resulted in a conformational change in the protein. Although HSP73 prevents aggregation of unfolded rhodanese in vitro, the chaperone activity of HSP73 was suppressed in the presence of GM. Using limited proteolysis of HSP73 by TPCK-trypsin, the GM binding site is a COOH-terminal for one third of the protein known to be a peptide-binding domain. During immunohistochemistry, HSP73 and GM were co-localized in enlarged lysosomes of rat kidneys with GM-induced acute tubular injury in vivo. Our results suggest that the specific association between HSP73 and GM may reduce the chaperone activity of HSP73 in vitro and/or in vivo, and this may have an interaction with GM toxicity in kidneys with GM-induced acute tubular injury.

Gut epithelial inducible heat-shock proteins and their modulation by diet and the microbiota

“…82 The latter effect may have resulted from the gentamicin-specific inhibition of HSP70's chaperoning activity, thus promoting a positive feedback response. 83,84 In normal pigs, oral administration of the broad-spectrum antibiotic amoxicillin to mothers around parturition was found to affect the mother's fecal and the offspring's gut microbiota as well as the offspring's gut epithelial iHSPs: small intestinal iHSP27 remained unaffected, while iHSP70 decreased transiently, and both iHSP proteins increased in colonic mucosa. 45,63 Data were interpreted as being essentially related to amoxicillin-induced changes in maternal microbiota and subsequent microbiota transfer to offspring after birth.…”

Arabidopsis Cor15am Is a Chloroplast Stromal Protein That Has Cryoprotective Activity and Forms Oligomers

“…CD spectra were determined as described previously (Miyazaki et al, 2004) for 60 mM recombinant Cor15am-His in 50 mM HEPES-KOH buffer, pH 7.5, using a J-720 spectropolarimeter (JUSCO). For some experiments, the protein sample was either boiled or subjected to a freeze-thaw treatment before the measurement.…”

Increased expression of co-chaperone HOP with HSP90 and HSC70 and complex formation in human colonic carcinoma

“…HSP90 associates with steroid hormone receptors (e.g., glucocorticoid receptor, androgen receptor, and estrogen receptor) and regulates the transport and maturation of the receptors (Carrigan et al 2005;Fiskus et al 2007). We have shown that the HSP70 and HSP90 chaperones strongly interact with a number of medical drugs (Ishida et al 2008;Miyazaki et al 2004;Otaka et al 2007).…”