Chemical and Local Effects on Peptide Cysteine Reactivity
Poster Number
16a
Introduction/Abstract
When a disulfide bond (R-S-S-R’) forms between the thiol groups of two cysteine amino acids this residue is known as cystine. Cysteine and cystine play a major role in protein structure. Disulfide bonds are usually formed in organisms by enzymes. The disulfide linkages can be scrambled in a protein or peptide with multiple disulfide bonds. This is associated with changes in protein structure and aggregation. Non-enzymatic disulfide formation presents challenges when studying cysteine containing peptides and proteins in the laboratory.
Purpose
Our current research involves controlled studies on disulfide bond formation and measurements of the relative acidities of simple cysteine containing peptides.
Method
All peptides are synthesized by solid phase synthesis using Rink amide resin. The kinetics of disulfide bond formation are monitored by using mass spectrometry technique. All peptides are measured in reference to the analog internal standard peptides Ser-Ala-Ala-Ala-Ala and Ser-Ala-Ala-Ala-Ser-Ala-Ala-Ala. Methanol:water 1:1(v/v) is used as the solvent for all reactions. Relative acidity is determined through titration. The peptide are titrated with sodium hydroxide and hydrochloric acid utilizing a micro pH probe.
Results
We have found that disulfide bonds are formed between small cysteine containing peptides in solution in the absence of enzymes at different rates. Proximity to the N-terminus of the peptide and the presence of metal ions including copper and magnesium accelerates disulfide formation. The presence of catalytic amounts of copper (II) causes an overall increase in the rate of disulfide bond formation in N- and C-terminal cysteine containing peptides, while the presence of magnesium (II) increases the rate of disulfide bond formation with an order of magnitude lesser effect than copper (II). Preliminary titration curves suggest the peptide Cys-Ala-Ala is more acidic than the rearranged peptide Ala-Ala-Cys.
Significance
This research generates insight into how peptide structure and properties influence the reactivity of cysteine residues. Cysteine and cystine play a major role in protein folding and the elimination of damaging oxidative free radicals in biological systems. Several neurodegenerative misfolding diseases including Alzheimer’s disease, Parkinson’s disease, prion-related disorders, and amyotrophic lateral sclerosis (ALS) are associated with disulfide bond scrambling. All pathways for the biological formation of disulfide bonds are not known. This research will facilitate the understanding of factors influencing disulfide bonds in peptides and proteins on the fundamental level.
Location
DeRosa University Center
Format
Poster Presentation
Poster Session
Morning 10am-12pm
Chemical and Local Effects on Peptide Cysteine Reactivity
DeRosa University Center
When a disulfide bond (R-S-S-R’) forms between the thiol groups of two cysteine amino acids this residue is known as cystine. Cysteine and cystine play a major role in protein structure. Disulfide bonds are usually formed in organisms by enzymes. The disulfide linkages can be scrambled in a protein or peptide with multiple disulfide bonds. This is associated with changes in protein structure and aggregation. Non-enzymatic disulfide formation presents challenges when studying cysteine containing peptides and proteins in the laboratory.
