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Zeolite molecular sieves have dramatic acid-base effects on enzymes in nonaqueous media

Fontes, N. and Partridge, J. and Halling, P.J. and Barreiros, S. (2002) Zeolite molecular sieves have dramatic acid-base effects on enzymes in nonaqueous media. Biotechnology and Bioengineering, 77 (3). pp. 296-305. ISSN 0006-3592

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Zeolite molecular sieves very commonly are used as in situ drying agents in reaction mixtures of enzymes in nonaqueous media. They often affect enzyme behavior, and this has been interpreted in terms of altered hydration. Here, we show that zeolites can also have dramatic acid-base effects on enzymes in low water media, resulting from their nation-exchange ability. Initial rates of transesterification catalyzed by crosslinked crystals of subtilisin were compared in supercritical ethane, hexane, and acetonitrile with water activity fixed by pre- equilibration. Addition of zeolite NaA (4 Angstrom powder) still caused remarkable rate enhancements (up to 20-fold), despite the separate control of hydration. In the presence of excess of an alternative solid-state acid-base buffer, however, zeolite addition had no effect. The more commonly used Merck molecular sieves (type 3 A beads) had similar but somewhat smaller effects. All zeolites have ion-exchange ability and can exchange H+ for cations such as Na+ and K+. These exchanges will tend to affect the protonation state of acidic groups in the protein and, hence, enzymatic activity. Zeolites pre- equilibrated in aqueous suspensions of varying pH-pNa gave very different enzyme activities. Their differing basicities were demonstrated directly by equilibration with an indicator dissolved in toluene. The potential of zeolites as acid-base buffers for low-water media is discussed, and their ability to overcome pH memory is demonstrated.