Determination of membrane protein glycation in diabetic tissue
Journal Title: The AAPS Journal - Year 1999, Vol 1, Issue 4
Abstract
Diabetes-associated hyperglycemia causes glycation of proteins at reactive amino groups, which can adversely affect protein function Although the effects of glycation on soluble proteins are well characterized, there is no information regarding membrane-associated proteins, mainly because of the lack of reproducible methods to determine protein glycation in vivo. The current study was conducted to establish such a method and to compare the glycation levels of membrane-associated proteins derived from normal and diabetic tissue. We present a detailed sample preparation protocol based on the borohydride-periodate assay, modified to allow manipulation of animal tissue. Assay noise associated with extraction protocols and nonproteinaceous buffer components was eliminated by the using 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS) as a membrane detergent, applying desalting columns, and including a protein precipitation step. The glycation level of membrane proteins from diabetic rats is elevated to 4.89 nmol/mg protein (standard deviation [SD] 0.48) compared with normoglycemic control tissue (2.23 nmol/mg protein, SD 0.64). This result is consistent with and correlated to the total glycated hemoglobin levels in diabetic and normoglycemic rats. Using<100 μg protein, the described methods allow further study of protein glycation effects on the function of individual transporter proteins and the role of these modifications in diabetes.
Authors and Affiliations
Eric Y. Zhang, Peter W. Swaan
Keywords
Related Articles
- EP ID EP682126
- DOI 10.1208/ps010420
- Views 73
- Downloads 0
Assessing the cost-effectiveness of pharmacogenomics
The use of pharmacogenomics to individualize drug therapy offers the potential to improve drug effectiveness, reduce adverse side effects, and provide cost-effective pharmaceutical care. However, the combinations of dise...
Permeability Comparison between Hepatocyte and Low Efflux MDCKII Cell Monolayer
Determination of passive permeability is not only important for predicting oral absorption and brain penetration, but also for accurately predicting hepatic clearance. High throughput (HT) measurement of passive permeabi...
Summary Workshop Report: Bioequivalence, Biopharmaceutics Classification System, and Beyond
The workshop “Bioequivalence, Biopharmaceutics Classification System, and Beyond” was held May 21–23, 2007 in North Bethesda, MD, USA. This workshop provided an opportunity for pharmaceutical scie...
2012 AAPS National Biotech Conference Open Forum: A Perspective on the Current State of Immunogenicity Prediction and Risk Management
The immunogenicity profile of a biotherapeutic is determined by multiple product-, process- or manufacturing-, patient- and treatment-related factors and the bioanalytical methodology used to monitor for immunogenicity....
Effect of rise in simulated inspiratory flow rate and carrier particle size on powder emptying from dry powder inhalers
The purpose of this study was to evaluate the effect of carrier particle size and simulated inspiratory flow increase rate on emptying from dry powder inhalers (DPIs). Several flow rate ramps were created using a compute...