Abstract
Lyophilized enzyme powder is often used in organic solvents. However, the enzymatic activity decreases during the reaction process. In the present study, the relation between structural stability and enzymatic activity in an organic solvent was investigated. 13C cross-polarization magic angle spinning NMR spectroscopy was used to determine the secondary structure of lyophilized papain in the solid-state. Deconvolution of the peaks of the backbone carbonyl carbons suggested that the proportion of β-sheet conformation increased after lyophilization from a phosphate buffer solution. The esterification of N-benzyloxycarbonyl phenylalanylalanine amide was attempted using the lyophilized papain as a catalyst in anhydrous 1-propanol. The yield of ester was 46.1% after 48 h at 50°C, but this reaction slowed remarkably after 48 h. When the lyophilized papain was suspended in anhydrous 1-propanol for 7 days without the substrate, the proportion of β-sheet conformation was further increased and the suspended papain had no activity. These results suggest that the increase in β-sheet conformation caused inactivation of papain. The increase in β-sheet conformation caused by both lyophilization and suspension in propanol was found, which was related to a decrease in enzymatic activity.
| Original language | English |
|---|---|
| Pages (from-to) | 928-933 |
| Number of pages | 6 |
| Journal | Biotechnology and Bioengineering |
| Volume | 93 |
| Issue number | 5 |
| DOIs | |
| Publication status | Published - 2006 Apr 5 |
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Keywords
- CPMAS solid-state NMR
- Lyophilization
- Organic solvent
- Papain
- Structural stability
ASJC Scopus subject areas
- Biotechnology
- Microbiology
Cite this
Stability of protease in organic solvent : Structural identification by solid-state NMR of lyophilized papain before and after 1-propanol treatment and the corresponding enzymatic activities. / Matsubara, Teruhiko; Fujita, Risa; Sugiyama, Shigeru; Kawashiro, Katsuhiro.
In: Biotechnology and Bioengineering, Vol. 93, No. 5, 05.04.2006, p. 928-933.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Stability of protease in organic solvent
T2 - Structural identification by solid-state NMR of lyophilized papain before and after 1-propanol treatment and the corresponding enzymatic activities
AU - Matsubara, Teruhiko
AU - Fujita, Risa
AU - Sugiyama, Shigeru
AU - Kawashiro, Katsuhiro
PY - 2006/4/5
Y1 - 2006/4/5
N2 - Lyophilized enzyme powder is often used in organic solvents. However, the enzymatic activity decreases during the reaction process. In the present study, the relation between structural stability and enzymatic activity in an organic solvent was investigated. 13C cross-polarization magic angle spinning NMR spectroscopy was used to determine the secondary structure of lyophilized papain in the solid-state. Deconvolution of the peaks of the backbone carbonyl carbons suggested that the proportion of β-sheet conformation increased after lyophilization from a phosphate buffer solution. The esterification of N-benzyloxycarbonyl phenylalanylalanine amide was attempted using the lyophilized papain as a catalyst in anhydrous 1-propanol. The yield of ester was 46.1% after 48 h at 50°C, but this reaction slowed remarkably after 48 h. When the lyophilized papain was suspended in anhydrous 1-propanol for 7 days without the substrate, the proportion of β-sheet conformation was further increased and the suspended papain had no activity. These results suggest that the increase in β-sheet conformation caused inactivation of papain. The increase in β-sheet conformation caused by both lyophilization and suspension in propanol was found, which was related to a decrease in enzymatic activity.
AB - Lyophilized enzyme powder is often used in organic solvents. However, the enzymatic activity decreases during the reaction process. In the present study, the relation between structural stability and enzymatic activity in an organic solvent was investigated. 13C cross-polarization magic angle spinning NMR spectroscopy was used to determine the secondary structure of lyophilized papain in the solid-state. Deconvolution of the peaks of the backbone carbonyl carbons suggested that the proportion of β-sheet conformation increased after lyophilization from a phosphate buffer solution. The esterification of N-benzyloxycarbonyl phenylalanylalanine amide was attempted using the lyophilized papain as a catalyst in anhydrous 1-propanol. The yield of ester was 46.1% after 48 h at 50°C, but this reaction slowed remarkably after 48 h. When the lyophilized papain was suspended in anhydrous 1-propanol for 7 days without the substrate, the proportion of β-sheet conformation was further increased and the suspended papain had no activity. These results suggest that the increase in β-sheet conformation caused inactivation of papain. The increase in β-sheet conformation caused by both lyophilization and suspension in propanol was found, which was related to a decrease in enzymatic activity.
KW - CPMAS solid-state NMR
KW - Lyophilization
KW - Organic solvent
KW - Papain
KW - Structural stability
UR - http://www.scopus.com/inward/record.url?scp=33646041627&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=33646041627&partnerID=8YFLogxK
U2 - 10.1002/bit.20791
DO - 10.1002/bit.20791
M3 - Article
C2 - 16329143
AN - SCOPUS:33646041627
VL - 93
SP - 928
EP - 933
JO - Biotechnology and Bioengineering
JF - Biotechnology and Bioengineering
SN - 0006-3592
IS - 5
ER -