Identification and Quantitative Studies of Protein Immobilization Sites by Stable Isotope Labeling and Mass Spectrometry

Wa, Chunling; Cerny, Ronald L.; Hage, David S.

doi:10.1021/ac0609935

Cited by 29 publications

(39 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although free amine groups are involved in both the Schiff base immobilization method and glycation, these processes tend to involve different residues on HSA, making the resulting protein support a good model for the soluble forms of normal HSA or glycated HSA [26][27][28][29][30][31][32][33]48]. A control support was prepared in the same manner, but with no protein being added during the immobilization step.…”

Section: Preparation Of Supports and Columnsmentioning

confidence: 99%

Analysis of glipizide binding to normal and glycated human serum albumin by high-performance affinity chromatography

et al. 2015

Self Cite

View full text Add to dashboard Cite

In diabetes, the elevated levels of glucose in the bloodstream can result in the nonenzymatic glycation of proteins such as human serum albumin (HSA). This type of modification has been shown to affect the interactions of some drugs with HSA, including several sulfonylurea drugs that are used to treat type II diabetes. This study used high-performance affinity chromatography (HPAC) to examine the interactions of glipizide (i.e., a second-generation sulfonylurea drug) with normal HSA or HSA that contained various levels of in vitro glycation. Frontal analysis indicated that glipizide was interacting with both normal and glycated HSA through two general groups of sites: a set of relatively strong interactions and a set of weaker interactions with average association equilibrium constants at pH 7.4 and 37 °C in the range of 2.4-6.0 × 10(5) and 1.7-3.7 × 10(4) M(-1), respectively. Zonal elution competition studies revealed that glipizide was interacting at both Sudlow sites I and II, which were estimated to have affinities of 3.2-3.9 × 10(5) and 1.1-1.4 × 10(4) M(-1). Allosteric effects were also noted to occur for this drug between the tamoxifen site and the binding of R-warfarin at Sudlow site I. Up to an 18% decrease in the affinity for glipizide was observed at Sudlow site I ongoing from normal HSA to glycated HSA, while up to a 27% increase was noted at Sudlow site II. This information should be useful in indicating how HPAC can be used to investigate other drugs that have complex interactions with proteins. These results should also be valuable in providing a better understanding of how glycation may affect drug-protein interactions and the serum transport of drugs such as glipizide during diabetes.

show abstract

Section: Preparation Of Supports and Columnsmentioning

confidence: 99%

Analysis of glipizide binding to normal and glycated human serum albumin by high-performance affinity chromatography

et al. 2015

Self Cite

View full text Add to dashboard Cite

show abstract

“…Supports containing normal HSA or glycated HSA were immobilized to this support through the Schiff base method [56]. A previous study that determined which amine groups on HSA are involved in the Schiff base method has found that these residues tend to be different from those that are involved in glycation [57]. Control columns were prepared in the same manner but with no protein being added during the immobilization step.…”

Section: Methodsmentioning

confidence: 99%

Analysis of multi-site drug–protein interactions by high-performance affinity chromatography: Binding by glimepiride to normal or glycated human serum albumin

Matsuda

Zhao

Zheng

et al. 2015

Journal of Chromatography A

Self Cite

View full text Add to dashboard Cite

High-performance affinity chromatography (HPAC) was used in a variety of formats to examine multi-site interactions between glimepiride, a third-generation sulfonylurea drug, and normal or in vitro glycated forms of the transport protein human serum albumin (HSA). Frontal analysis revealed that glimepiride interacts with normal HSA and glycated HSA at a group of high affinity sites (association equilibrium constant, or K a , 9.2-11.8 × 10 5 M −1 at pH 7.4 and 37°C) and a group of lower affinity regions (K a , 5.9-16.2 × 10 3 M −1 ). Zonal elution competition studies were designed and carried out in both normal-and reversed-role formats to investigate the binding by this drug at specific sites. These experiments indicated that glimepiride was interacting at both Sudlow sites I and II. Allosteric effects were also noted with R-warfarin at Sudlow site I and with tamoxifen at the tamoxifen site on HSA. The binding at Sudlow site I had a 2.1-to 2.3-fold increase in affinity in going from normal HSA to the glycated samples of HSA. There was no significant change in the affinity for glimepiride at Sudlow site II in going from normal HSA to a moderately glycated sample of HSA, but a slight decrease in affinity was seen in going to a more highly glycated HSA sample. These results demonstrated how various HPAC-based methods can be used to profile and characterize multi-site binding by a drug such as glimepiride to a protein and its modified forms. The information obtained from this study should be useful in providing a better understanding of how drug-protein binding may be affected by glycation and of how separation and analysis methods based on HPAC can be employed to study systems with complex interactions or that involve modified proteins.

show abstract

“…As noted in prior work, the immobilization of HSA by this approach mainly occurs through the N -terminus or lysines that are not located at Sudlow sites I and II [37] and has been shown to provide columns with normal HSA or glycated HSA that are good models for the soluble forms of these proteins [10,18,21,22]. Control supports were prepared in the same manner but with no protein being added during the immobilization step.…”

Section: Methodsmentioning

confidence: 99%

Chromatographic analysis of the effects of fatty acids and glycation on binding by probes for Sudlow sites I and II to human serum albumin

Anguizola

DeBolt

Suresh

et al. 2016

Journal of Chromatography B

Self Cite

View full text Add to dashboard Cite

The primary endogenous ligands of human serum albumin (HSA) are non-esterified fatty acids, with 0.1–2 moles of fatty acids normally being bound to HSA. In type II diabetes, fatty acid levels in serum are often elevated, and the presence of high glucose results in an increase in the non-enzymatic glycation of HSA. High-performance affinity chromatography (HPAC) was used to examine the combined effects of glycation and the presence of long chain fatty acids on the binding of HSA with R-warfarin and L-tryptophan (i.e., probes for Sudlow sites I and II, the major sites for drugs on this protein). Zonal elution competition studies were used to examine the interactions of myristic acid, palmitic acid and stearic acid with these probes on HSA. It was found that all these fatty acids had direct competition with R-warfarin at Sudlow site I of normal HSA and glycated HSA, with the glycated HSA typically having stronger binding for the fatty acids at this site. At Sudlow site II, direct competition was observed for all the fatty acids with L-tryptophan when using normal HSA, while glycated HSA gave no competition or positive allosteric interactions between these fatty acids and L-tryptophan. These data indicated that glycation can alter the interactions of drugs and fatty acids at specific binding sites on HSA. The results of this study should lead to a better understanding of how these interactions may change during diabetes and demonstrate how HPAC can be used to examine drug/solute-protein interactions in complex systems.

show abstract

Identification and Quantitative Studies of Protein Immobilization Sites by Stable Isotope Labeling and Mass Spectrometry

Cited by 29 publications

References 42 publications

Analysis of glipizide binding to normal and glycated human serum albumin by high-performance affinity chromatography

Analysis of glipizide binding to normal and glycated human serum albumin by high-performance affinity chromatography

Analysis of multi-site drug–protein interactions by high-performance affinity chromatography: Binding by glimepiride to normal or glycated human serum albumin

Chromatographic analysis of the effects of fatty acids and glycation on binding by probes for Sudlow sites I and II to human serum albumin

Contact Info

Product

Resources

About