Non-enzymatic glycation induces structural modifications of myoglobin.
Increased glucose concentration in diabetes mellitus causes glycation of several proteins, leading to changes in their properties. Although glycation-induced functional modification of myoglobin is known, structural modification of the protein has not yet been reported. Here, we have studied glucose-modified structural changes of the heme protein. After in vitro glycation of metmyoglobin (Mb) by glucose at 25 degrees C for 6 days, glycated myoglobin (GMb) and unchanged Mb have been separated by ion exchange (BioRex 70) chromatography, and their properties have been compared. Compared to Mb, GMb exhibits increased absorbance around 280 nm and enhanced fluorescence emission with excitation at 285 nm. Fluorescence quenching experiments of the proteins by acrylamide and KI indicate that more surface accessible tryptophan residues are exposed in GMb. CD spectroscopic study reveals a change in the secondary structure of GMb with decreased alpha-helix content. 1-anilino-naphthaline-8-sulfonate (ANS) binding with Mb and GMb indicates that glycation increases hydrophobicity of the heme protein. GMb appears to be less stable with respect to thermal denaturation and differential calorimetry experiments. Heme-globin linkage becomes weaker in GMb, as shown by spectroscopic and gel electrophoresis experiments. A correlation between glycation-induced structural and functional modifications of the heme protein has been suggested.
Roy
Anjana
A
Department of Biophysics, Molecular Biology and Bioinformatics, University of Calcutta, Kolkata, India.
Sil
Rajarshi
R
Chakraborti
Abhay Sankar
AS
D000818:Animals; D002417:Cattle; D002942:Circular Dichroism; D005456:Fluorescent Dyes / Q000378:metabolism; D005947:Glucose / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D006736:Horses; D009211:Myoglobin / Q000737:chemistry* / Q000378:metabolism; D011489:Protein Denaturation; D055550:Protein Stability; D017433:Protein Structure, Secondary; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism
D005456:Fluorescent Dyes; D009211:Myoglobin; D014364:Tryptophan; D005947:Glucose