Evaluation of denaturation and aggregate formation of enzyme under heat stress conditions.

Kuboi, Ryoichi; Yamahara, Katsuhito; Ota, Hideyuki

doi:10.1252/jcej.30.1119

Cited by 18 publications

(10 citation statements)

References 8 publications

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“…One is a fraction which is detected from both A 280 and dA 420 /dt (numbers [19][20][21][22]. The other fractions are detected by only A 280 (numbers [23][24][25][26]. In both fractions of free -gal, the -gal could be detected in the silver-stained image of SDS-polyacrylamido gel electrophoresis (data not shown).…”

Section: Resultsmentioning

confidence: 99%

“…Edwards et al (33) have previously indicated the change of -gal conformation and the existence of inactive tetramer of -gal during the thermal denaturation process. The surface properties (e.g., surface net hydrophobicity, HFS, and local hydrophobicity, LH) have previously reported to be caused by such conformational change and analyzed by the aqueous two-phase partitioning method (18,25). The surface properties of -gal (HFS G and LH G ) were analyzed after the -gal was exposed to heat treatment.…”

Section: Variation Of Surface Properties Of Liposomes and -Gal Variamentioning

confidence: 99%

“…Such intermediate of -gal is thought to be formed during the thermal inactivation of -gal (Figure 7). It has been reported that the LH values of carbonic anhydrase (from bovine, CAB) having various conformations were investigated, and the local hydrophobicity, LH, values of CAB of the partial unfolded states indicated relatively high values (18,25,37). In the case of -gal, the LH G value of the intermediate states is much higher than that of other states ( Figure 8), and the -gal of such state is, therefore, thought to interact easily with the hydrophobic binding sites of liposome surface ( Figure 6).…”

Section: Variation Of Surface Properties Of Liposomes and -Gal Variamentioning

confidence: 99%

“…Recently, heat-induced translocation of cytoplasmic -galactosidase ( -gal) across the inner membrane of Escherichia coli has been investigated (20)(21)(22)(23). The plausible scheme for heat-induced translocation has been shown based on the evaluated properties of protein surface (24,25) and cell membrane (26,27) under the stress condition, where the possible roles of GroEL and other chaperons have also been discussed (23). The phospholipid membrane has been reported to assist the refolding of denatured protein through the hydrophobic interaction to the protein of the partly unfolded states (28).…”

Section: Introductionmentioning

confidence: 99%

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Model System for Heat‐Induced Translocation of Cytoplasmic β‐Galactosidase across Phospholipid Bilayer Membrane

Umakoshi

Yoshimoto

Shimanouchi

et al. 1998

Biotechnology Progress

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The possibility of the translocation of the enzyme across the phospholipid bilayer membrane was investigated by using the liposomes prepared by 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) in which beta-galactosidase (beta-gal) was entrapped. Exposing the POPC liposomes entrapping beta-gal inside to heat treatment (40-50 degrees C, 1-60 min) was found to induce its translocation across the liposome membrane. The translocated activity of beta-gal from inner to outer aqueous phase of liposomes indicated the maximal value when the liposomes entrapping beta-gal were heated at 45 degrees C for 30 min. The gel permeation profiles of the liposomes before and after heat treatment (45 degrees C, 30 min) also supported the translocation of beta-gal across the liposome membrane. The membrane fluidity of liposomes was found to be increased with increasing temperature, so that the hydrophobicity of liposome membrane was also increased. The local hydrophobicity of beta-gal was maximized at the temperature of 40-50 degrees C. The mechanisms of beta-gal translocation have been suggested to be triggered by the enhancement of hydrophobic interaction between the liposome surface and beta-gal molecules. Finally, a minimal scheme of possible mechanism on the heat-induced translocation of beta-gal has been presented on the basis of the hydrophobic interaction between the liposome and the proteins. The experimental data on the heat-induced translocation of beta-gal were well corresponding to those from model calculation.

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Section: Resultsmentioning

confidence: 99%

Section: Variation Of Surface Properties Of Liposomes and -Gal Variamentioning

confidence: 99%

Section: Variation Of Surface Properties Of Liposomes and -Gal Variamentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Model System for Heat‐Induced Translocation of Cytoplasmic β‐Galactosidase across Phospholipid Bilayer Membrane

Umakoshi

Yoshimoto

Shimanouchi

et al. 1998

Biotechnology Progress

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“…There are several reports noting that conformational changes of proteins could be monitored by measurements of the amounts of adsorption of detergents, such as 8-anilinonaphthalene-1-sulfonate (ANS) and Triton X-100 (TX), on protein surfaces in solution (Semistonov et al, 1991;Kuboi et al, 1997). Nonionic detergents such as Triton X series were reported to be adsorbed to proteins without profound effects on protein conformation (Makino et al, 1973) and to interact with the hydrophobic binding sites of proteins (Border, 1981).…”

Section: Introductionmentioning

confidence: 99%

Measuring Adsorption of a Hydrophobic Probe with a Surface Plasmon Resonance Sensor to Monitor Conformational Changes in Immobilized Proteins

Yamaguchi

Mannen²,

Zako

et al. 2008

Biotechnol Progress

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Conformational changes of proteins immobilized on solid matrices were observed by measuring the adsorption of Triton X-100 (TX), a nonionic detergent, as a hydrophobic probe with BIACORE, a biosensor that utilizes the phenomenon of surface plasmon resonance (SPR). Two kinds of proteins, alpha-glucosidase and lysozyme, were covalently attached to dextran matrices on the sensor surface in the flow cell and then exposed to various concentrations of TX solution. We measured SPR signal changes derived from adsorption of TX to the immobilized proteins and calculated the monolayer adsorption capacity using the Brunauer-Emmett-Teller (BET) equation. The results demonstrated that monolayer adsorption capacity is proportional to the amount of immobilized proteins. Further, the unfolding process of immobilized proteins on the sensor surface induced by guanidine hydrochloride was investigated by monitoring SPR signal increases due to the adsorption of TX to the exposed hydrophobic region of the protein. Results strongly suggested that the increase in the SPR signal reflected the formation of the agglutinative unfolded state. We expect our measuring method using the SPR sensor and TX adsorption will be a novel tool to provide conformational information regarding various proteins on solid matrices.

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Study on Stress‐Mediated Behavior and Preparation of Giant Vesicles

Shimanouchi

Umakoshi

Kuboi

1999

Perspectives in Supramolecular Chemistry

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Evaluation of denaturation and aggregate formation of enzyme under heat stress conditions.

Cited by 18 publications

References 8 publications

Model System for Heat‐Induced Translocation of Cytoplasmic β‐Galactosidase across Phospholipid Bilayer Membrane

Model System for Heat‐Induced Translocation of Cytoplasmic β‐Galactosidase across Phospholipid Bilayer Membrane

Measuring Adsorption of a Hydrophobic Probe with a Surface Plasmon Resonance Sensor to Monitor Conformational Changes in Immobilized Proteins

Study on Stress‐Mediated Behavior and Preparation of Giant Vesicles

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