Control of enzyme ionization state in supercritical ethane by sodium/proton solid-state acid-base buffers

N. Fontes, P.J. Halling, S. Barreiros

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

We have previously demonstrated that a solid-state buffer could be successfully used to control the ionization state of subtilisin Carlsberg cross-linked microcrystals (CLECs) suspended in supercritical ethane (sc-ethane) in the presence of acid-base active species such as salt hydrates and zeolite molecular sieves. Here we studied the effect of six zwitterionic proton/sodium (pH-pNa) solid-state acid-base buffers on the catalytic activity of subtilisin CLECs in sc- ethane at high and low water activity (a(W)). CLECs were strongly activated by increasing a(W). At high a(W), and despite the high hydrolysis rates, transesterification activities were still about one order of magnitude higher than those observed at lower a(W). This is in contradiction with what was previously reported in the absence of acid-base control and supports the hypothesis that the poor catalytic performance of subtilisin CLECs at high a(W) observed in those studies was due to the inhibitory effect of the hydrolytic by- product, rather than to the competition of water with propanol for the acyl-enzyme intermediate. Although the catalytic activity of subtilisin showed a general positive correlation with the aqueous pK(a) of the acid-base buffers tested here, our results also show that as expected, the acid-base behavior of the buffers in nonaqueous media is more complex than what can be predicted from aqueous-based parameters alone. This work further confirms the usefulness of solid-state acid-base buffers in supercritical biocatalysis but highlights the need for further research on the topic.
LanguageEnglish
Pages938-941
Number of pages3
JournalEnzyme and Microbial Technology
Volume33
Issue number7
DOIs
Publication statusPublished - 2 Dec 2003

Fingerprint

Ethane
Ionization
Protons
Buffers
Microcrystals
Sodium
Subtilisin
Acids
Enzymes
Catalyst activity
Biocatalysis
Subtilisins
Zeolites
1-Propanol
Water
Transesterification
Molecular sieves
Hydrates
Byproducts
Hydrolysis

Keywords

  • enzymes
  • supercritical fluids
  • water activity
  • enzyme ionization
  • solid-state buffers Organic media
  • nonaqueous media
  • catalysis
  • solvents
  • biocatalysis
  • crystals
  • fluids

Cite this

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title = "Control of enzyme ionization state in supercritical ethane by sodium/proton solid-state acid-base buffers",
abstract = "We have previously demonstrated that a solid-state buffer could be successfully used to control the ionization state of subtilisin Carlsberg cross-linked microcrystals (CLECs) suspended in supercritical ethane (sc-ethane) in the presence of acid-base active species such as salt hydrates and zeolite molecular sieves. Here we studied the effect of six zwitterionic proton/sodium (pH-pNa) solid-state acid-base buffers on the catalytic activity of subtilisin CLECs in sc- ethane at high and low water activity (a(W)). CLECs were strongly activated by increasing a(W). At high a(W), and despite the high hydrolysis rates, transesterification activities were still about one order of magnitude higher than those observed at lower a(W). This is in contradiction with what was previously reported in the absence of acid-base control and supports the hypothesis that the poor catalytic performance of subtilisin CLECs at high a(W) observed in those studies was due to the inhibitory effect of the hydrolytic by- product, rather than to the competition of water with propanol for the acyl-enzyme intermediate. Although the catalytic activity of subtilisin showed a general positive correlation with the aqueous pK(a) of the acid-base buffers tested here, our results also show that as expected, the acid-base behavior of the buffers in nonaqueous media is more complex than what can be predicted from aqueous-based parameters alone. This work further confirms the usefulness of solid-state acid-base buffers in supercritical biocatalysis but highlights the need for further research on the topic.",
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Control of enzyme ionization state in supercritical ethane by sodium/proton solid-state acid-base buffers. / Fontes, N.; Halling, P.J.; Barreiros, S.

In: Enzyme and Microbial Technology, Vol. 33, No. 7, 02.12.2003, p. 938-941.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Control of enzyme ionization state in supercritical ethane by sodium/proton solid-state acid-base buffers

AU - Fontes, N.

AU - Halling, P.J.

AU - Barreiros, S.

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AB - We have previously demonstrated that a solid-state buffer could be successfully used to control the ionization state of subtilisin Carlsberg cross-linked microcrystals (CLECs) suspended in supercritical ethane (sc-ethane) in the presence of acid-base active species such as salt hydrates and zeolite molecular sieves. Here we studied the effect of six zwitterionic proton/sodium (pH-pNa) solid-state acid-base buffers on the catalytic activity of subtilisin CLECs in sc- ethane at high and low water activity (a(W)). CLECs were strongly activated by increasing a(W). At high a(W), and despite the high hydrolysis rates, transesterification activities were still about one order of magnitude higher than those observed at lower a(W). This is in contradiction with what was previously reported in the absence of acid-base control and supports the hypothesis that the poor catalytic performance of subtilisin CLECs at high a(W) observed in those studies was due to the inhibitory effect of the hydrolytic by- product, rather than to the competition of water with propanol for the acyl-enzyme intermediate. Although the catalytic activity of subtilisin showed a general positive correlation with the aqueous pK(a) of the acid-base buffers tested here, our results also show that as expected, the acid-base behavior of the buffers in nonaqueous media is more complex than what can be predicted from aqueous-based parameters alone. This work further confirms the usefulness of solid-state acid-base buffers in supercritical biocatalysis but highlights the need for further research on the topic.

KW - enzymes

KW - supercritical fluids

KW - water activity

KW - enzyme ionization

KW - solid-state buffers Organic media

KW - nonaqueous media

KW - catalysis

KW - solvents

KW - biocatalysis

KW - crystals

KW - fluids

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DO - 10.1016/j.enzmictec.2003.07.003

M3 - Article

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EP - 941

JO - Enzyme and Microbial Technology

T2 - Enzyme and Microbial Technology

JF - Enzyme and Microbial Technology

SN - 0141-0229

IS - 7

ER -