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Role of the n+1 amino acid residue on the deamidation of asparagine in pentapeptides
Hasan H. Incea, F. Aylin Sungur Konuklarb, Ilke Ugurcde, Ö. Alaz Ozcand, Maryam Sayadif, Michael Feigg & Viktorya Aviyented
a Department of Chemistry, Michigan State University, East Lansing, USA
b Informatics Institute, Computational Science and Engineering Division, Istanbul Technical
University, Istanbul, Turkey
c Université de Lorraine, Vandoeuvre-les-Nancy, France
d Department of Chemistry, Bogaziçi University, Istanbul, Turkey
e CNRS, Vandoeuvre-les-Nancy, France
f Kim Lab, University of Toronto, Toronto, Canada
g Department of Chemistry and Department of Biochemistry & Molecular Biology, Michigan
State University, East Lansing, USA Published online: 27 Jul 2015.
To cite this article: Hasan H. Ince, F. Aylin Sungur Konuklar, Ilke Ugur, Ö. Alaz Ozcan, Maryam Sayadi, Michael Feig & Viktorya
Aviyente (2015): Role of the n+1 amino acid residue on the deamidation of asparagine in pentapeptides, Molecular Physics: An International Journal at the Interface Between Chemistry and Physics, DOI: 10.1080/00268976.2015.1068394
To link to this article: http://dx.doi.org/10.1080/00268976.2015.1068394
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Molecular Physics, 2015 http://dx.doi.org/10.1080/00268976.2015.1068394
RESEARCH ARTICLE
Role of the n + 1 amino acid residue on the deamidation of asparagine in pentapeptides
Hasan H. Incea , F. Aylin Sungur Konuklarb , Ilke Ugurc , d , e , O¨ . Alaz Ozcand , Maryam Sayadif , Michael Feigg and
Viktorya Aviyented , ‗
a Department of Chemistry, Michigan State University, East Lansing, USA; b Informatics Institute, Computational Science and
Engineering Division, Istanbul Technical University, Istanbul, Turkey; c Universite´ de Lorraine, Vandoeuvre-les-Nancy, France;
d Department of Chemistry, Bogazic¸i University, Istanbul, Turkey; e CNRS, Vandoeuvre-les-Nancy, France; f Kim Lab, University of Toronto, Toronto, Canada; g Department of Chemistry and Department of Biochemistry & Molecular Biology, Michigan State University, East Lansing, USA
(Received 29 March 2015; accepted 26 June 2015)
Deamidation plays an important role in biochemical phenomena such as aging. The role of the n + 1 residue on the deamidation of asparagine (asparagine being the nth residue) in three pentapeptide chains (GGNGG, GGNMG and GGNIG) has been analysed with hybrid computational tools. Potentials of mean force at 300 K were calculated from the MD/replica exchange simulations using weighted histogram analysis (WHAM) in explicit water. The snapshots were clustered taking into account the requirements of the plausible deamidation mechanisms, as such the tautomerisation of the asparagine side chain as initial step has been confirmed, based on the proximity of water to the deamidation site. The ultimate goal being to gain an insight on the peptide backbone N-H acidity, quantum mechanical calculations have been carried out. For this purpose, the distribution of / , 2 / and end-to-end distances deduced from the WHAM diagrams have been considered and a total of 110 structures have been sampled. These neutral pentapeptides as well as their corresponding anions have been
optimised (B3LYP/6-31 ++ G(d,p)) in implicit water in order to gain an insight on the peptide backbone N-H acidity. In this
study, we have shown that the open conformations of the neutrals and the anions, which display a β sheet like structure are
well populated and their pKa s rank in the same order as the deamidating half-lives, that is the peptides that deaminate fastest can more readily access conformations that are more acidic.
Keywords: deamidation; peptide; modelling
1. Introduction
Deamidation is the conversion of the amide group on the side chain of an amino acid residue to a carboxylate or car- boxylic acid depending on the pH of the medium. Under physiological conditions, proteins spontaneously degrade via physical and chemical processes. It was proposed that accumulation of protein defects, resulting from deamida- tion, may be one of the main causes of human aging [1]. Two of the twenty ordinary amino acid residues in peptides and proteins, asparagine (Asn) and glutamine (Gln), are un- stable under physiological conditions [2]. Deamidation of Asn or Gln introduces a negative charge at the deamidation residue, and thus the structure of the deamidating peptide or protein changes. The conversion of the neutral amide side chain to the negatively charged carboxylate causes time dependent changes in conformation and limits the life- time of peptides and proteins [3–6]. It has been suggested that sequence-dependent non-enzymatic amide hydrolysis, deamidation of glutaminyl and asparaginyl residues in pep- tides and proteins may serve as a general molecular timer of biological processes [7].
The non-enzymatic deamidation is also detected in pep- tides in vitro. The chemical instability introduced by this re- action in vitro causes several problems in pharmaceuticals. For instance, growth hormone-releasing factors were found to lose their biological activities because of being subject to deamidation during storage [8]. This major instability as- cribed to deamidation was observed years before the recent developments in engineered protein drugs. Currently, con- trolling the rate of deamidation is one of the major aspects of protein therapeutics optimisation, since it is one of the most commonly occurred degradations in peptides [9,10].
Under neutral and basic conditions, the deamidation is base catalysed and the products are a mixture of as- partate (Asp), and iso-aspartate (Iso-Asp). Production of Iso-Asp requires cleavage of the peptide backbone. The experimentally isolated cyclic imide intermediate (succin- imide) justifies the backbone contribution in basic and neu- tral media [11]. The deprotonation of the peptide bond nitrogen is necessary for succinimide-mediated deami- dation and isomerisation. Capasso et al. defined succin- imide formation as the rate determining step of the overall
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‗ Corresponding author. Email: aviye@boun.edu.tr
C 2015 Taylor & Francis
Scheme 1. Reactants and products of Asn deamidation.
reaction [12]. The cyclic imide then hydrolyses at either one of the two carbonyls to yield a mixture of Asp and Iso-Asp residues. The ratio of L-Asp to L-iso-Asp was experimen- tally found to be 3:1 (Scheme 1).
Tyler-Cross and Schirch [13] and Robinson et al. [1] found that the residue immediately following the Asn (on the C-terminal side) has a large effect on the deamidation rate. The fastest deamidation rates, on the time scale of a day at neutral pH, were observed for sequences containing - Asn-Gly- [1]. Kosky et al. [14], Xie et al. [15] and Robinson et al. [16–18] found that secondary and tertiary structures generally reduce deamidation rates as compared to peptides with the same sequence. Peters and Trout claim that the conversion of Asn to succinimide is the rate limiting step in the deamidation [19].
The most rapid deamidation was observed when the
n + 1 residue is glycine (Gly). The facilitating effect of Gly
on deamidation with respect to other residues was explained
by several factors:
(1) Steric hindrance: The size of groups on the α- carbon of n + 1 residue is seen to control the rate
of deamidation. However, the degradation rates are not linearly correlated with the size of the residues [20,21].
(2) Backbone amide acidity: Experimental studies have related the deamidation rate to the hydrogen ex- change rate constants of the backbone amide hy-
drogen of the n + 1 residue and have created a new
perspective to the facilitating effect of Gly [22]. The
assumption here was that the rate of deamidation is directly correlated to acidity of backbone amide hy-
drogen of n + 1. This assumption fairly explained the impact of n + 1 residue for several non- Gly
residues. However, the relative rate of the peptides
wit...