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The anionic biosurfactant rhamnolipid does not denature industrial enzymes.

Madsen JK, Pihl R, Møller AH, Madsen AT, Otzen DE, Andersen KK - Front Microbiol (2015)

Bottom Line: It efficiently unfolds both LT and CZ, but LT is unfolded by SDS through formation of SDS clusters on the enzyme well below the cmc, while CZ is only unfolded by bulk micelles and on average binds significantly less SDS than LT.In contrast, RL does not affect the tertiary or secondary structure of any enzyme at room temperature, has little impact on thermal stability and only binds detectably (but at low stoichiometries) to SZ.Furthermore, all enzymes maintain activity at both monomeric and micellar concentrations of RL.

View Article: PubMed Central - PubMed

Affiliation: Interdisciplinary Nanoscience Center (iNANO), Department of Molecular Biology and Genetics, Aarhus University Aarhus, Denmark.

ABSTRACT
Biosurfactants (BS) are surface-active molecules produced by microorganisms. Their combination of useful properties and sustainable production make them promising industrial alternatives to petrochemical and oleochemical surfactants. Here we compare the impact of the anionic BS rhamnolipid (RL) and the conventional/synthetic anionic surfactant sodium dodecyl sulfate (SDS) on the structure and stability of three different commercially used enzymes, namely the cellulase Carezyme® (CZ), the phospholipase Lecitase Ultra® (LT) and the α-amylase Stainzyme® (SZ). Our data reveal a fundamental difference in their mode of interaction. SDS shows great diversity of interaction toward the different enzymes. It efficiently unfolds both LT and CZ, but LT is unfolded by SDS through formation of SDS clusters on the enzyme well below the cmc, while CZ is only unfolded by bulk micelles and on average binds significantly less SDS than LT. SDS binds with even lower stoichiometry to SZ and leads to an increase in thermal stability. In contrast, RL does not affect the tertiary or secondary structure of any enzyme at room temperature, has little impact on thermal stability and only binds detectably (but at low stoichiometries) to SZ. Furthermore, all enzymes maintain activity at both monomeric and micellar concentrations of RL. We conclude that RL, despite its anionic charge, is a surfactant that does not compromise the structural integrity of industrially relevant enzymes. This makes RL a promising alternative to current synthetic anionic surfactants in a wide range of commercial applications.

No MeSH data available.


Related in: MedlinePlus

Using pyrene fluorescence to determine SDS and RL cmc and cluster formation in the presence of the three enzymes. (A) SDS: The I3/I1 ratio of pyrene changes around 2–3 mM SDS in the absence of enzymes. In the presence of SZ and LT the I3/I1 ratio already starts to rise at 0.25 and 1 mM SDS, respectively, indicating formation of SDS micellar clusters on the enzyme surface below the cmc. (B) RL: The I3/I1 ratio of pyrene changes around 0.1–1 mM RL in the absence of enzymes. The increase of the I3/I1 ratio increases already at 0.05 mM in the presence of SZ suggests that RL forms micellar clusters on the surface of SZ below the cmc.
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Figure 1: Using pyrene fluorescence to determine SDS and RL cmc and cluster formation in the presence of the three enzymes. (A) SDS: The I3/I1 ratio of pyrene changes around 2–3 mM SDS in the absence of enzymes. In the presence of SZ and LT the I3/I1 ratio already starts to rise at 0.25 and 1 mM SDS, respectively, indicating formation of SDS micellar clusters on the enzyme surface below the cmc. (B) RL: The I3/I1 ratio of pyrene changes around 0.1–1 mM RL in the absence of enzymes. The increase of the I3/I1 ratio increases already at 0.05 mM in the presence of SZ suggests that RL forms micellar clusters on the surface of SZ below the cmc.

Mentions: The cmc of SDS is around 7 mM in water but reduces with increasing ionic strength (Jönsson et al., 1998). Incubation of pyrene with increasing concentrations of SDS reveals a systematic development in the I3/I1 fluorescence ratio with increasing SDS concentration: the ratio is stable around 0.6 between 0 and ~2 mM SDS where after it increases to reach a plateau of 0.9 at ~3 mM SDS (Figure 1A). This indicates that micelles are formed in solution around 2–3 mM SDS. The three enzymes all behaved in different ways in the pyrene model system. CZ did not change the titration pattern (Figure 1A), indicating that SDS does not form micellar structures on CZ below the cmc. In contrast, both LT and SZ lead to change in pyrene fluorescence below the cmc. For SZ the I3/I1 ratio increased from 0.6 to a plateau around 0.7 already at 0.25 mM SDS, indicating interactions at very low SDS concentrations. The ratio then merged with the protein-free SDS curve around the cmc. For LT the ratio was stable at 0.6 until ~0.75 mM SDS where after the I3/I1 ratio steadily increased with increasing SDS concentration, but only merged with the protein-free sample at a ratio of ~0.8, in the middle of the transition region.


The anionic biosurfactant rhamnolipid does not denature industrial enzymes.

Madsen JK, Pihl R, Møller AH, Madsen AT, Otzen DE, Andersen KK - Front Microbiol (2015)

Using pyrene fluorescence to determine SDS and RL cmc and cluster formation in the presence of the three enzymes. (A) SDS: The I3/I1 ratio of pyrene changes around 2–3 mM SDS in the absence of enzymes. In the presence of SZ and LT the I3/I1 ratio already starts to rise at 0.25 and 1 mM SDS, respectively, indicating formation of SDS micellar clusters on the enzyme surface below the cmc. (B) RL: The I3/I1 ratio of pyrene changes around 0.1–1 mM RL in the absence of enzymes. The increase of the I3/I1 ratio increases already at 0.05 mM in the presence of SZ suggests that RL forms micellar clusters on the surface of SZ below the cmc.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400916&req=5

Figure 1: Using pyrene fluorescence to determine SDS and RL cmc and cluster formation in the presence of the three enzymes. (A) SDS: The I3/I1 ratio of pyrene changes around 2–3 mM SDS in the absence of enzymes. In the presence of SZ and LT the I3/I1 ratio already starts to rise at 0.25 and 1 mM SDS, respectively, indicating formation of SDS micellar clusters on the enzyme surface below the cmc. (B) RL: The I3/I1 ratio of pyrene changes around 0.1–1 mM RL in the absence of enzymes. The increase of the I3/I1 ratio increases already at 0.05 mM in the presence of SZ suggests that RL forms micellar clusters on the surface of SZ below the cmc.
Mentions: The cmc of SDS is around 7 mM in water but reduces with increasing ionic strength (Jönsson et al., 1998). Incubation of pyrene with increasing concentrations of SDS reveals a systematic development in the I3/I1 fluorescence ratio with increasing SDS concentration: the ratio is stable around 0.6 between 0 and ~2 mM SDS where after it increases to reach a plateau of 0.9 at ~3 mM SDS (Figure 1A). This indicates that micelles are formed in solution around 2–3 mM SDS. The three enzymes all behaved in different ways in the pyrene model system. CZ did not change the titration pattern (Figure 1A), indicating that SDS does not form micellar structures on CZ below the cmc. In contrast, both LT and SZ lead to change in pyrene fluorescence below the cmc. For SZ the I3/I1 ratio increased from 0.6 to a plateau around 0.7 already at 0.25 mM SDS, indicating interactions at very low SDS concentrations. The ratio then merged with the protein-free SDS curve around the cmc. For LT the ratio was stable at 0.6 until ~0.75 mM SDS where after the I3/I1 ratio steadily increased with increasing SDS concentration, but only merged with the protein-free sample at a ratio of ~0.8, in the middle of the transition region.

Bottom Line: It efficiently unfolds both LT and CZ, but LT is unfolded by SDS through formation of SDS clusters on the enzyme well below the cmc, while CZ is only unfolded by bulk micelles and on average binds significantly less SDS than LT.In contrast, RL does not affect the tertiary or secondary structure of any enzyme at room temperature, has little impact on thermal stability and only binds detectably (but at low stoichiometries) to SZ.Furthermore, all enzymes maintain activity at both monomeric and micellar concentrations of RL.

View Article: PubMed Central - PubMed

Affiliation: Interdisciplinary Nanoscience Center (iNANO), Department of Molecular Biology and Genetics, Aarhus University Aarhus, Denmark.

ABSTRACT
Biosurfactants (BS) are surface-active molecules produced by microorganisms. Their combination of useful properties and sustainable production make them promising industrial alternatives to petrochemical and oleochemical surfactants. Here we compare the impact of the anionic BS rhamnolipid (RL) and the conventional/synthetic anionic surfactant sodium dodecyl sulfate (SDS) on the structure and stability of three different commercially used enzymes, namely the cellulase Carezyme® (CZ), the phospholipase Lecitase Ultra® (LT) and the α-amylase Stainzyme® (SZ). Our data reveal a fundamental difference in their mode of interaction. SDS shows great diversity of interaction toward the different enzymes. It efficiently unfolds both LT and CZ, but LT is unfolded by SDS through formation of SDS clusters on the enzyme well below the cmc, while CZ is only unfolded by bulk micelles and on average binds significantly less SDS than LT. SDS binds with even lower stoichiometry to SZ and leads to an increase in thermal stability. In contrast, RL does not affect the tertiary or secondary structure of any enzyme at room temperature, has little impact on thermal stability and only binds detectably (but at low stoichiometries) to SZ. Furthermore, all enzymes maintain activity at both monomeric and micellar concentrations of RL. We conclude that RL, despite its anionic charge, is a surfactant that does not compromise the structural integrity of industrially relevant enzymes. This makes RL a promising alternative to current synthetic anionic surfactants in a wide range of commercial applications.

No MeSH data available.


Related in: MedlinePlus