Reverse vesicle formation from the yeast glycolipid biosurfactant mannosylerythritol lipid-D.

Mannosylerythritol lipids (MELs) are secreted by yeasts and are promising glycolipid biosurfactants. In our study on the non-aqueous phase behaviors of MEL homologues, we found that MEL-D (4-O-[2',3'-di-O-alka(e)noyl-ß-D-mannopyranosyl]-(2R,3S)-erythritol) forms aggregates in decane. The microscopic observation and the X-ray scattering measurement of these aggregates revealed that they are reverse vesicles that consist of bilayers whose hydrophilic domains are located in the interior of the bilayers. In addition, MEL-D formed reverse vesicles without co-surfactants and co-solvents in various oily solutions, such as n-alkanes, cyclohexane, squalane, squalene, and silicone oils at a concentration below 10 mM. This is the first report on the reverse vesicle formation from biosurfactants.

Packing density of glycolipid biosurfactant monolayers give a significant effect on their binding affinity toward immunoglobulin G.

Mannosylerythritol lipid-A (MEL-A) is one of the most promising glycolipid biosurfactants, and abundantly produced by Pseudozyma yeasts. MEL-A gives not only excellent self-assembling properties but also a high binding affinity toward human immunoglobulin G (HIgG). In this study, three kinds of MEL-A were prepared from methyl myristate [MEL-A (m)], olive oil [MEL-A (o)], and soybean oil [MEL-A (s)], and the effect of interfacial properties of each MEL-A monolayer on the binding affinity toward HIgG was investigated using surface plasmon resonance (SPR) and the measurement of surface pressure (pi)-area (A) isotherms. Based on GC-MS analysis, the main fatty acids were C(8) and C(10) acids in all MEL-A, and the content of unsaturated fatty acids was 0% for MEL-A (m), 9.1% for MEL-A (o), 46.3% for MEL-A (s), respectively. Interestingly, the acid content significantly influenced on their binding affinity, and the monolayer of MEL-A (o) gave a higher binding affinity than that of MEL-A (m) and MEL-A (s). Moreover, the mixed MEL-A (o)/ MEL-A (s) monolayer prepared from 1/1 molar ratio, which comprised of 27.8% of unsaturated fatty acids, indicated the highest binding affinity. At the air/water interface, MEL-A (o) monolayer exhibited a phase transition at 13 degrees C from a liquid condensed monolayer to a liquid expanded monolayer, and the area per molecule significantly expanded above 13 degrees C, while the amount of HIgG bound to the liquid expanded monolayer was much higher than that bound to liquid condensed monolayer. The binding affinity of MEL-A toward HIgG is thus likely to closely relate to the monolayer packing density, and may be partly controlled by temperature.

Kinetic studies on the interactions between glycolipid biosurfactant assembled monolayers and various classes of immunoglobulins using surface plasmon resonance.

Kinetic studies on the interactions between self-assembled monolayers of mannosylerythritol lipids (MELs), which are glycolipid biosurfactants abundantly produced by microorganisms, and various classes of immunoglobulins including human IgG, IgA, and IgM were performed using surface plasmon resonance (SPR). The effect of the MEL structure on the binding behavior of HIgG was examined. Assembled monolayers of MEL-A having two acetyl groups on the headgroup gave a high affinity (K(d)=1.7x10(-6)M) toward HIgG, while those of MEL-B or MEL-C having only one acetyl group at C-6' or C-4' position gave little affinity. Our kinetic analysis revealed that the binding manner of HIgG, HIgA (K(d)=2.4x10(-7)M), and HIgM (K(d)=2.2x10(-7)M) to the assembled monolayers of MEL-A is not the monovalent mode but the bivalent mode, and both the first and second rate association constants (k(a1), k(a2)) increase with an increase in the number of antibody binding sites, while those for dissociation (k(d1), k(d2)) changed little. Moreover, we succeeded in directly observing great amounts of HIgG, HIgA, and HIgM bound to MEL-A monolayers using atomic force microscopy (AFM). Finally, we found that MEL-A assembled monolayer binds toward various IgG derived from mouse, pig, rabbit, horse, goat, rat, and bovine as well as human IgG (HIgG), and the only exception was sheep IgG. These results clearly demonstrate that MEL-A assembled monolayers would be useful as noble affinity ligand system for various immunoglobulins.

Monolayers assembled from a glycolipid biosurfactant from Pseudozyma (Candida) antarctica serve as a high-affinity ligand system for immunoglobulin G and M.

A carbohydrate ligand system has been developed which is composed of self-assembled monolayers (SAMs) of mannosylerythritol lipid-A (MEL-A) from Pseudozyma antarctica, serving for human immunoglobulin G and M (HIgG and HIgM). The estimated binding constants from surface plasmon resonance (SPR) measurement were Ka = 9.4 x 10(6) M(-1) for HIgG and 5.4 x 10(6) M(-1) for HIgM, respectively. The binding site was not in the Fc region of immunoglobulin but in the Fab region. Large amounts of HIgG and HIgM bound to MEL-A SAMs were directly observed by atomic force microscopy.