Deamidation effect

S. J. Wearne and T. E. Creighton, Effect of protein conformation on rate of deamidation Ribonuclease A, Proteins Struct. Funct. Genet, 5, 8 (1989). 

R. Tyler-Cross and V. Schirch, Effects of amino acid sequence, buffers and ionic strength on the rate and mechanism of deamidation of asparagine residues in small peptides, J. Biol. Chem, 266, 22549 (1991). 

Lai, M.C., Hageman, M.J., Schowen, R.L., Borchardt, R.T., and Topp, E.M. 1999a. Chemical stability of peptides in polymers. 1. Effect of water on peptide deamidation in poly(vinyl alcohol) and poly(vinyl pyrrolidone) matrixes. J. Pharm. Sci. 88, 1073-1080. 

The influence of secondary structure on reactions of deamidation has been confirmed in a number of studies. Thus, deamidation was inversely proportional to the extent of a-helicity in model peptides [120], Similarly, a-hel-ices and /3-turns were found to stabilize asparagine residues against deamidation, whereas the effect of /3-sheets was unclear [114], The tertiary structure of proteins is also a major determinant of chemical stability, in particular against deamidation [121], on the basis of several factors such as the stabilization of elements of secondary structure and restrictions to local flexibility, as also discussed for the reactivity of aspartic acid residues (Sect. 6.3.3). Furthermore, deamidation is markedly decreased in regions of low polarity in the interior of proteins because the formation of cyclic imides (Fig. 6.29, Pathway e) is favored by deprotonation of the nucleophilic backbone N-atom, which is markedly reduced in solvents of low polarity [100][112],... 

An example of this effect is provided by ribonuclease A (RNase A). At pH 8 and 37°, the rate of deamidation of Asn67 was more than 30-fold lower in the native than in the unfolded protein [111]. Deamidation of the native RNase A was also ca. 30-fold slower than of an octapeptide whose sequence is similar to that of the deamidation site, although the reaction mechanisms were similar [108][123],... 

In neutral and alkaline solutions, the iso-Asp/Asp-peptide products were always formed in a ca. 3 1 ratio. In the Val-Xaa-Asn-Ser-Val series, the nature of the N-flanking residue had little effect on the rate of deamidation, regardless of size. However, the actual effect may have been partly masked by cleavage of the Asn-Ser bond (Sect. 6.3.2A), which proceeds at a rate ca. 10% that of deamidation. In the Val-Ser-Asn-Xaa-Val series, the nature of the C-flanking residue did have a major influence. As expected, the most unstable peptide was Val-Ser-Asn-Gly-Val (tll2 ca. 6 h at pH 7.3 and 60°). A second group of peptides had t1/2 values in the range of 25-75 h, which increased in the order Xaa=His[Pg.325]

C. Goolcharran, L. L. Stauffer, J. L. Cleland, R. T. Borchardt, The Effects of a Histidine Residue on the C-Terminal Side of an Asparaginyl Residue on the Rate of Deamidation Using Model Pentapeptides , J. Pharm. Sci. 2000, 89, 818-825. 

S. Capasso, G. Balboni, P. Di Cerbo, Effect of Lysine Residues on the Deamidation Reaction of Asparagine Side-Chains , Biopolymers 2000, 53, 213 - 219. 

It has been shown in a number of instances that under identical conditions the reaction rates of amorphous forms are greater than in crystalline forms of the same dmg [21]. Generally, water has a destabilising effect in the majority of cases, for example, deamidation, hydrolysis or oxidation [27]. 

Whether there is any other connection between anticonvulsant activity and camosine s antiaging actions is obviously highly speculative. It may be relevant to note that epileptic seizures and a shortened life span, together with altered protein accumulation, are consequences of PIMT-deficiency in mice, while treatment with valproic acid, an anticonvulsant, partially suppresses these symptoms including effects on life span (Yamamoto et ah, 1998). Conversely, PIMT overexpression can increase life span of Drosophila (Bennet et ah, 2003). Furthermore, the chemistry of some anticonvulsants (ethosuximide) resembles quite closely the structure of the succinimide intermediate formed during both asparagine deamidation and PIMT-mediated repair of isoaspartate residues. One conjectures whether there are any relationships between these... 

The present study was conducted to obtain additional information on changes in soy protein subunits during limited proteolysis. Enzymatic soy protein deamidation that occurred, in addition to limited proteolysis, during germination of soybean seeds was investigated. The effects of proteolysis and deamidation on solubility and emulsifying activity were compared. Phosphorylation of soy protein with a commercially available protein kinase and its effects on subsequent changes in functional properties of the protein were also studied. 

Emulsifying activity index (EAI) is a measure of the ability of protein to emulsify oil, which depends on solubility, size, charge, and surface activity of the protein molecules. The effect of proteolysis with pronase E on EAI of the modified protein was relatively insignificant (Figure 6) However, deamidation appeared to enhance EAI, especially at pH values more basic than the isoelectric point (pH 4.7). 

In limited proteolysis, proteases such as pronase E hydrolyzed the 7S subunits of soy proteins more than the IIS subunits, resulting in enhanced protein solubility. Deamidation with relatively insignificant peptide bond hydrolysis that occurred during the germination of soybeans imparted to the storage protein improved solubility and emulsifying activity. On the other hand, the incorporation of phosphorus in soy proteins by the protein kinase cAMPdPK was too low to effect significant... 

Two examples where metal ions confer stability or increased activity in proteins are human deoxyribonuclease (rhDNase, Pulmozyme ), and Factor VIII. In the case of rhDNase, Ca2+ ions (up to 100 mM) increased the stability of the enzyme through a specific binding site (64). In fact, the removal of calcium ions from the solution with EGTA caused an increase in deamidation and aggregation. However, this effect was observed only with Ca+2 ions other divalent cations, Mg2+, Mn2+, and Zn2+, were observed to destabilize rhDNase. Similar effects were observed in Factor VIII. Ca2+ and Sr2+ ions stabilized the protein, whereas others such as Mg2+, Mn2+ and Zn2+, Cu2+, and Fe2+ destabilized the enzyme (65). In a separate study with Factor VIII, a significant increase in the aggregation rate was observed in the presence of Al3+ ions (66). The authors note that other excipients like buffer salts are often contaminated with Al3+ ions and illustrate the need to use excipients of appropriate quality in formulated products. 

Patel K, Borchardt RT. Chemical pathways of peptide degradation. III. Effect of primary sequence on the pathways of deamidation of asparaginyl residues in hexapeptides. Pharm Res 1990 7(8) 787-793. 

---