The effect of the presence of globular proteins and elongated polymers on enzyme activity.

We have studied the effect of a crowded (macromolecular) solution on reaction rates of the decarboxylating enzymes urease, pyruvate decarboxylase and glutamate decarboxylase. A variety of crowding agents were used including haemoglobin, lysozyme, various dextrans and polyethylene glycol. Enzyme reaction rates of all three enzymes show two different types of effect that separate the globular proteins from the polysaccharides/polymers. Increasing concentration of globular proteins caused a dramatic rise in enzyme activity up to 30% crowding concentration then the activity decreased, whereas the polymers caused a concentration dependent decrease in activity. The viscosities of the globular proteins were low compared to the polymers. The increased activity with proteins may be due to the decreased amount of free water, which leads to higher effective concentration of substrates, or to an increased oligomeric state by self-association. The lower activities of all enzymes with all agents at high concentrations may be explained by a decrease in rates of diffusion. An increase in protein crowding (decrease in cell water content) may contribute to pathological conditions including cataract and Alzheimer's disease.

[1]  A. Minton.  Implications of macromolecular crowding for protein assembly. , 2000, Current opinion in structural biology.

[2]  R. Ellis,et al.  Macromolecular crowding: an important but neglected aspect of the intracellular environment. , 2001 .

[3]  E. Bakker,et al.  Transient accumulation of potassium glutamate and its replacement by trehalose during adaptation of growing cells of Escherichia coli K-12 to elevated sodium chloride concentrations , 2004, Archives of Microbiology.

[4]  A. Minton,et al.  Macromolecular Crowding Accelerates Amyloid Formation by Human Apolipoprotein C-II* , 2002, The Journal of Biological Chemistry.

[5]  B. Derham,et al.  Enzyme activity after resealing within ghost erythrocyte cells, and protection by alpha-crystallin against fructose-induced inactivation. , 2002, The Biochemical journal.

[6]  A. Bron,et al.  Physical properties of stimulated and unstimulated tears. , 1999, Experimental eye research.

[7]  T. Schindler,et al.  Diffusion control in an elementary protein folding reaction. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[8]  C. Slingsby,et al.  Rapid separation of bovine -crystallin subunits B1, B2, B3, A3 and A4 , 1990 .

[9]  S. Zimmerman,et al.  Macromolecular crowding increases binding of DNA polymerase to DNA: an adaptive effect. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[10]  J. J. Harding,et al.  Cataract: Biochemistry, Epidemiology and Pharmacology , 1991 .

[11]  R. Lindner,et al.  Effects of dextran on the self-association of human spectrin. , 1995, Biophysical chemistry.

[12]  M. Record,et al.  Responses of E. coli to osmotic stress: large changes in amounts of cytoplasmic solutes and water. , 1998, Trends in biochemical sciences.

[13]  A. Minton,et al.  Macromolecular crowding: biochemical, biophysical, and physiological consequences. , 1993, Annual review of biophysics and biomolecular structure.

[14]  D J Jenden,et al.  In vivo proton magnetic resonance spectroscopy of the normal aging human brain. , 1996, Life sciences.

[15]  Kiowa S. Bower,et al.  Accelerated α‐synuclein fibrillation in crowded milieu , 2002 .

[16]  G. Ralston,et al.  Enhancement of self-association of human spectrin by polyethylene glycol. , 1994, The International journal of biochemistry.

[17]  A. Minton.  Excluded volume as a determinant of macromolecular structure and reactivity , 1981 .

[18]  A. Minton,et al.  Effect of macromolecular crowding upon the structure and function of an enzyme: glyceraldehyde-3-phosphate dehydrogenase. , 1981, Biochemistry.

[19]  S. Zimmerman,et al.  Estimation of macromolecule concentrations and excluded volume effects for the cytoplasm of Escherichia coli. , 1991, Journal of molecular biology.