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Journal of Bacteriology, August 2002, p. 4384-4391, Vol. 184, No. 16
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.16.4384-4391.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Trehalose-Mediated Inhibition of the Plasma Membrane H+-ATPase from Kluyveromyces lactis: Dependence on Viscosity and Temperature

José G. Sampedro,1* Rosario A. Muñoz-Clares,2 and Salvador Uribe1

Departamento de Bioquímica, Instituto de Fisiología Celular,1 Departamento de Bioquímica, Facultad de Química, Universidad Nacional Autónoma de México, 04510 Mexico City, México2

Received 7 February 2002/ Accepted 31 May 2002

The effect of increasing trehalose concentrations on the kinetics of the plasma membrane H+-ATPase from Kluyveromyces lactis was studied at different temperatures. At 20°C, increasing concentrations of trehalose (0.2 to 0.8 M) decreased Vmax and increased S0.5 (substrate concentration when initial velocity equals 0.5 Vmax), mainly at high trehalose concentrations (0.6 to 0.8 M). The quotient Vmax/S0.5 decreased from 5.76 µmol of ATP mg of protein-1 min-1 mM-1 in the absence of trehalose to 1.63 µmol of ATP mg of protein-1 min-1 mM-1 in the presence of 0.8 M trehalose. The decrease in Vmax was linearly dependent on solution viscosity ({eta}), suggesting that inhibition was due to hindering of protein domain diffusional motion during catalysis and in accordance with Kramer's theory for reactions in solution. In this regard, two other viscosity-increasing agents, sucrose and glycerol, behaved similarly, exhibiting the same viscosity-enzyme inhibition correlation predicted. In the absence of trehalose, increasing the temperature up to 40°C resulted in an exponential increase in Vmax and a decrease in enzyme cooperativity (n), while S0.5 was not modified. As temperature increased, the effect of trehalose on Vmax decreased to become negligible at 40°C, in good correlation with the temperature-mediated decrease in viscosity. The trehalose-mediated increase in S0.5 was similar at all temperatures tested, and thus, trehalose effects on Vmax/S0.5 were always observed. Trehalose increased the activation energy for ATP hydrolysis. Trehalose-mediated inhibition of enzymes may explain why yeast rapidly hydrolyzes trehalose when exiting heat shock.

* Corresponding author. Mailing address: Departamento de Bioquímica, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México (UNAM), A.P. 70-242, 04510 Mexico City, DF, México. Phone: (5255)-5622-5632. Fax: (5255)-5622-5630. E-mail: jsamped{at}ifisiol.unam.mx.

Journal of Bacteriology, August 2002, p. 4384-4391, Vol. 184, No. 16
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.16.4384-4391.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Conlin, L. K., Nelson, H. C. M. (2007). The Natural Osmolyte Trehalose Is a Positive Regulator of the Heat-Induced Activity of Yeast Heat Shock Transcription Factor. Mol. Cell. Biol. 27: 1505-1515 [Abstract] [Full Text]  


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