The importance of amphipathic (amphoteric) polymers in biological transport has been recognized for many years. Only recently have recombinant DNA techniques been employed to produce or isolate well defined, repetitive polypeptide sequences and to study their phase transfer, sequestration, and transport roles. These techniques allow for precise control of polymer microstructure and also extend the possibility of incorporating predetermined secondary structure into the resulting polymers.
While biosynthetic technologies have experienced remarkable growth in a short period of time, most applications have been limited to the fields of medicine, pharmacy, or agriculture. A recent symposium sponsored by the American Chemical Society, however, has revealed a small but growing number of interdisciplinary efforts in materials development.
In the past several years, we have became interested in the exchangeable apolipoprotein, ApoLpIII from the insect Locusta migratoria. Exchangeable apolipoproteins are defined as those apolipoproteins capable of moving from one lipoprotein particle to another, versus the non-exchangeable apolipoproteins that remain with one lipoprotein particle from biosynthesis to catabolism. Lipoprotein particles in the insect hemolymph result from the interaction of the principle apolipoprotein particle high-density lipophorin (HDLp) with lipids. During lipid loading, ApoLpIII, existing as a free monomer, associates with the HDLp particle to form a low-density lipophorin (LDLp). Upon hydrolysis the LDLp, HDLp and ApoLpIII are released into the hemolymph and may be reused to load and shuttle lipid at the fat body.
This protein is of interest as a result of its ability to reversibly interact with the surface of a lipoprotein particle. the behavior of ApoLpIII has been attributed to various secondary structural elements present. the protein is composed of five amphipathic a-helices, with opposing polar and non-polar faces aligned along the long axis of the helix. thus these helices possess distinctly hydrophobic faces. In this proteins, regions that behave as "hinges" have been postulated to facilitate a rearrangement of the apolipoprotein secondary structure elements in order to regulate the protein composition of lipoprotein particles. Upon rearrangement, the hydrophobic portions of the individual helices interact with the hydrophobic surface of the lipoprotein particle. Such a naturally occurring protein apparently utilizes hydrophobic interactions to reversibly associate with hydrophobic lipoprotein particles in the aqueous environment.
Currently our laboratories have synthesized ApoLpIII recombinantly and are currently characterizing the protein's ability to complex with hydrophobic substances. Characterization methods include dynamic and classical light scattering experiments to determine size and mobility of protein-hydrophobe complexes, fluorescence measurements (both steady-state and time-resolved) to determine polarity of microenvironments within the protein, and determining the changes of tertiary structure with changes in environmental conditions.