An attempt to modulate the microporous diffusion of a model polypeptide by altering its secondary structure.
The purpose of the present study was to determine whether intentional alteration of the secondary structure of a model polypeptide, conantokin-G, influenced the rate and extent of aqueous pore diffusion across a synthetic microporous membrane. Use of a microporous synthetic membrane allowed for analysis of polypeptide transport without the confounding variables of protein binding, acid- and/or enzyme-mediated degradation, endocytotic uptake, and enzymatic inactivation associated with a biological membrane. Conantokin-G was intentionally changed from its native random coil structure to the alpha-helix structure using calcium, and both structures were verified using circular dichroism. The alpha-helix structure of conantokin-G was retained even after additional free calcium was removed by equilibrium dialysis. Over the concentration range of 1.25 to 20 mM, there was a linear relationship between the solution calcium concentration and the percent of the alpha-helix conformer present. The apparent permeability, the apparent aqueous diffusion coefficient with and without inclusion of the Renkin function, and the hydrodynamic radii estimated by diffusion and a computer-software program were calculated for the random coil and alpha-helix structures of conantokin-G. Calcium-mediated conversion of conantokin-G to its alpha-helix structure did not significantly (p >.05) change its apparent permeability across a microporous membrane. It is suggested that perhaps complete conversion to the alpha-helix structure of only a fraction of the conantokin-G molecules (only 0.45 or 45% of the molecules can be converted to the alpha-helix structure at Ca(2+) concentrations >or= 20 mM) may have limited the extent of transport of the alpha-helix conformer.
Johnston
Thomas P
TP
Division of Pharmaceutical Sciences, School of Pharmacy, University of Missouri-Kansas City, Missouri 64110-2499, USA. johnstont@umkc.edu
Salamat-Miller
Nazila T
NT
Alur
Hemant H
HH
Mitra
Ashim K
AK
D000595:Amino Acid Sequence; D000818:Animals; D002626:Chemistry, Pharmaceutical / Q000379:methods*; D020916:Conotoxins / Q000737:chemistry* / Q000378:metabolism; D004058:Diffusion / Q000187:drug effects*; D004304:Dosage Forms; D008567:Membranes, Artificial; D008851:Micropore Filters; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D010539:Permeability / Q000187:drug effects; D017433:Protein Structure, Secondary; D012908:Snails; D013329:Structure-Activity Relationship
D020916:Conotoxins; D004304:Dosage Forms; D008567:Membranes, Artificial; D010455:Peptides; C055407:conotoxin GV