Estimating conformation content of a protein using citrate-stabilized Au nanoparticles.
Herein we report the use of the optical properties of citrate-stabilized gold nanoparticles (Au NPs) for estimation of native or denatured conformation content in a mixture of a protein in solution. The UV-vis extinction spectrum of citrate-stabilized Au NPs is known to broaden differently in the presence of native and denatured states of alpha-amylase, bovine serum albumin (BSA) or amyloglucosidase (AMG). On the other hand, herein we show that when a mixture of native and denatured protein was present in the medium, the broadening of the spectrum differed for different fractional content of the conformations. Also, the total area under the extinction spectrum varied linearly with the change in the mole fraction content of a state and for a constant total protein concentration. Transmission electron microscopy (TEM) measurements revealed different levels of agglomeration for different fractional contents of the native or denatured state of a protein. In addition, time-dependent denaturation of a protein could be followed using the present method. The rate constants calculated for denaturation indicated a possible fast change in conformation of a protein before complete thermal denaturation. The observations have been explained based on the changes in extinction coefficient (thereby oscillator strength) upon interaction of citrate-stabilized NPs with proteins being in different states and levels of agglomeration.
Deka
Jashmini
J
Department of Chemistry, Indian Institute of Technology, Guwahati, 781 039, India.
Paul
Anumita
A
Chattopadhyay
Arun
A
D000818:Animals; D002417:Cattle; D019343:Citric Acid / Q000737:chemistry*; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry; D006046:Gold / Q000737:chemistry*; D053768:Metal Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D011489:Protein Denaturation; D011506:Proteins / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013056:Spectrophotometry, Ultraviolet; D013696:Temperature; D000516:alpha-Amylases / Q000737:chemistry
D011506:Proteins; D012710:Serum Albumin, Bovine; D019343:Citric Acid; D006046:Gold; D000516:alpha-Amylases; D005087:Glucan 1,4-alpha-Glucosidase