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Rosin Surfactant QRMAE Can Be Utilized as an Amorphous Aggregate Inducer: A Case Study of Mammalian Serum Albumin.

Ishtikhar M, Chandel TI, Ahmad A, Ali MS, Al-Lohadan HA, Atta AM, Khan RH - PLoS ONE (2015)

Bottom Line: It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI.Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate.Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins.

View Article: PubMed Central - PubMed

Affiliation: Protein Biophysics Laboratory, Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh - 202002, India.

ABSTRACT
Quaternary amine of diethylaminoethyl rosin ester (QRMAE), chemically synthesized biocompatible rosin based cationic surfactant, has various biological applications including its use as a food product additive. In this study, we examined the amorphous aggregation behavior of mammalian serum albumins at pH 7.5, i.e., two units above their isoelectric points (pI ~5.5), and the roles played by positive charge and hydrophobicity of exogenously added rosin surfactant QRMAE. The study was carried out on five mammalian serum albumins, using various spectroscopic methods, dye binding assay, circular dichroism and electron microscopy. The thermodynamics of the binding of mammalian serum albumins to cationic rosin modified surfactant were established using isothermal titration calorimetry (ITC). It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI. Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate. Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins. This study can be used for the design of novel biomolecules or drugs with the ability to neutralize factor(s) responsible for the aggregate formation, in addition to various other industrial applications.

No MeSH data available.


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(A) RLS measurements of the mammalian serum albumins samples at 350 nm were obtained in the absence and presence of varying concentrations of QRMAE to find out the maximum aggregation points at which surfactant concentration maximum aggregate was formed. (B) RLS measurement of protein samples in the absence and presence of rosin surfactant QRMAE at pH below and above two unit of their pI. Albumin concentration was fixed 5 μM in all conditions. The experiment was performed after 60 min incubation at 65°C.
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pone.0139027.g002: (A) RLS measurements of the mammalian serum albumins samples at 350 nm were obtained in the absence and presence of varying concentrations of QRMAE to find out the maximum aggregation points at which surfactant concentration maximum aggregate was formed. (B) RLS measurement of protein samples in the absence and presence of rosin surfactant QRMAE at pH below and above two unit of their pI. Albumin concentration was fixed 5 μM in all conditions. The experiment was performed after 60 min incubation at 65°C.

Mentions: The aggregation propensity of QRMAE at different concentrations was once again analyzed by RLS measurements at 350 nm with the all five mammalian albumins in the absence and presence of rosin surfactant at pH two unit above and below of pI. This technique is highly sensitive in determining the aggregation or turbidity of proteins. The increase in fluorescence intensity at 350 nm of protein samples incubated with QRMAE at pH above two units of its pI revealed the extent of scattering of light in the protein. We found that the maximum intensity was revealed at 250 μM concentration of QRMAE, only at pH above two unit of pI (Fig 2A). This information showed an excellent uniformity of the result obtained by the turbidity experiments. Furthermore, at that particular concentration of QRMAE, surfactant was incubated with all five types of albumins at two particular pH values i.e. two units above and below its pI. A noteworthy increase in fluorescence intensity (approximately several fold) was observed when proteins were incubated with QRMAE in buffer above two units of its pI at temperature 65°C, as compared to control samples (Fig 2B), The values are shown in Table 1. The turbidity measurement was highly supported by RLS results. Maximum absorption obtained by turbidity measurement and maximum fluorescence intensity shown by RLS analysis was found to be at the same concentration of the surfactant (Figs 1A and 2A). Thus, this ensures that the formation of aggregates occurs in all the five serum albumin samples at particular surfactant concentration and pH value.


Rosin Surfactant QRMAE Can Be Utilized as an Amorphous Aggregate Inducer: A Case Study of Mammalian Serum Albumin.

Ishtikhar M, Chandel TI, Ahmad A, Ali MS, Al-Lohadan HA, Atta AM, Khan RH - PLoS ONE (2015)

(A) RLS measurements of the mammalian serum albumins samples at 350 nm were obtained in the absence and presence of varying concentrations of QRMAE to find out the maximum aggregation points at which surfactant concentration maximum aggregate was formed. (B) RLS measurement of protein samples in the absence and presence of rosin surfactant QRMAE at pH below and above two unit of their pI. Albumin concentration was fixed 5 μM in all conditions. The experiment was performed after 60 min incubation at 65°C.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4587963&req=5

pone.0139027.g002: (A) RLS measurements of the mammalian serum albumins samples at 350 nm were obtained in the absence and presence of varying concentrations of QRMAE to find out the maximum aggregation points at which surfactant concentration maximum aggregate was formed. (B) RLS measurement of protein samples in the absence and presence of rosin surfactant QRMAE at pH below and above two unit of their pI. Albumin concentration was fixed 5 μM in all conditions. The experiment was performed after 60 min incubation at 65°C.
Mentions: The aggregation propensity of QRMAE at different concentrations was once again analyzed by RLS measurements at 350 nm with the all five mammalian albumins in the absence and presence of rosin surfactant at pH two unit above and below of pI. This technique is highly sensitive in determining the aggregation or turbidity of proteins. The increase in fluorescence intensity at 350 nm of protein samples incubated with QRMAE at pH above two units of its pI revealed the extent of scattering of light in the protein. We found that the maximum intensity was revealed at 250 μM concentration of QRMAE, only at pH above two unit of pI (Fig 2A). This information showed an excellent uniformity of the result obtained by the turbidity experiments. Furthermore, at that particular concentration of QRMAE, surfactant was incubated with all five types of albumins at two particular pH values i.e. two units above and below its pI. A noteworthy increase in fluorescence intensity (approximately several fold) was observed when proteins were incubated with QRMAE in buffer above two units of its pI at temperature 65°C, as compared to control samples (Fig 2B), The values are shown in Table 1. The turbidity measurement was highly supported by RLS results. Maximum absorption obtained by turbidity measurement and maximum fluorescence intensity shown by RLS analysis was found to be at the same concentration of the surfactant (Figs 1A and 2A). Thus, this ensures that the formation of aggregates occurs in all the five serum albumin samples at particular surfactant concentration and pH value.

Bottom Line: It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI.Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate.Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins.

View Article: PubMed Central - PubMed

Affiliation: Protein Biophysics Laboratory, Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh - 202002, India.

ABSTRACT
Quaternary amine of diethylaminoethyl rosin ester (QRMAE), chemically synthesized biocompatible rosin based cationic surfactant, has various biological applications including its use as a food product additive. In this study, we examined the amorphous aggregation behavior of mammalian serum albumins at pH 7.5, i.e., two units above their isoelectric points (pI ~5.5), and the roles played by positive charge and hydrophobicity of exogenously added rosin surfactant QRMAE. The study was carried out on five mammalian serum albumins, using various spectroscopic methods, dye binding assay, circular dichroism and electron microscopy. The thermodynamics of the binding of mammalian serum albumins to cationic rosin modified surfactant were established using isothermal titration calorimetry (ITC). It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI. Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate. Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins. This study can be used for the design of novel biomolecules or drugs with the ability to neutralize factor(s) responsible for the aggregate formation, in addition to various other industrial applications.

No MeSH data available.


Related in: MedlinePlus