Asn and Gin side chains

Reactions are carried out at room temperature in dichloromethane which is an excellent swelling agent for polystyrene supports. Dimethylformamide can be added to increase the solubility of some amino acid derivatives e.g. Boc-Arg(N02), Boc-Arg(Tos), Boc-His(Tos), Boc-Trp, and Boc-Asn(Xan), but it increases the rate of undesirable rearrangement of O-acylisourea to A-acylurea which is not reactive.P The principal limitations in using car-bodiimides are racemization, rearrangement of O-acylisourea to A-acylurea 5, and dehydration of Asn and Gin side-chain carboxamide groups. Fortunately this dehydration problem is completely avoided by the use of additives such as A-hydroxysuccinimide (HOSu) and 1,2,3-benzotriazol-l-ol (HOBt). Additives also reduce racemization and A-acyl-urea formation. Other carbodiimides have also been used in SPPS such as or N-tert-... 

Use of relaxation data at two magnetic fields to derive extended model-free parameters ps-ms dynamics of Asn and Gin side chains... 

To explore this issue, Monte Carlo computer simulations were run using the protocol outlined in the previous section. In these simulations, however, a peptide of sequence Ac-Ala-Xaa-Ala-Ala-NMe was employed (Xaa = Gin or Asn), the backbone was not constrained to the PPII conformation, and a side chain-to-backbone hydrogen bond was constrained using a potential function previously used to constrain a-helical backbone-to-backbone hydrogen bonds (Tun and Hermans, 1991 Creamer and Rose, 1994). 

Why should some peptide bonds be more sensitive to hydrolysis than others As discussed in the next section, the side chains of certain residues (e.g., Asp, Glu, Asn, and Gin) are able to undergo intramolecular reaction with the adjacent residue, leading to a variety of peptide bond fission and rearrangement reactions. Certain residues when located at the N- or C-termi-nus also exhibit a particular degree of reactivity. 

Figure 3-28 A15N - H HSQC spectrum of partially denatured 129-residue hen lysozyme. Boxes enclose the tryptophan indole region (upper left), the arginine side chain NE region (upper left), and a portion of the amide NH region (lower center and enlarged in the insert). Resonances of pairs of hydrogen atoms in side chain (Asn and Gin) amide groups are indicated by horizontal lines. From Buck et al.52i...

Figure 12-15 Schematic drawing of the active site of a cysteine protease of the papain family with a partial structure of an acyl-enzyme intermediate in green. The thiolate-imidazolium pair of Cys 25 His 159 lies deep in the substrate-binding cleft and bridges an interface between two major structural domains, just as the Ser His pair does in serine proteases (Fig. 12-10). This may facilitate small conformational changes during the catalytic cycle. Asn 175 provides a polarizable acceptor for positive charge, helping to stabilize the preformed ion pair, and allows easy transfer of an imidazolium proton to the product of substrate cleavage. The peptide NH of Cys 25 and the side chain of Gin 19 form an oxyanion hole.

An enzyme can deactivate irreversibly for two kinds of reasons (i) conformational processes, such as aggregation (intermolecular), or incorrect structure formation (intramolecular), such as scrambled disulfide bond formation between wrong side chains, and (ii) covalent processes, such as reduction and thus destruction of disulfide bonds, deamidation of asparagine (Asn) or glutamine (Gin) side chains, or hydrolysis of (usually) labile asp-X bonds in the protein sequence. 

Side-chains display a much stronger tendency to act as donors than as acceptors of hydrogen bonds. This is observed in particular for the O-H groups of Ser, Thr, and for the imidazole group of His. The residues Asn and Gin (in those cases where amide C=0 and NH2 could be distinguished), show no definite preference. 

Individual position substitutions on TMIII at Ilel24 and Leul27 give rise to receptors with constitutive activity (Baranski et al, 1999). When changed to Asn and Gin respectively, the mutant receptors signal in the absence of G5a. Since these positions are functionally conserved as hydro-phobic side chains in rhodopsin and the Oib and / 2 adrenoceptors, it would be interesting if analogous substitutions induced the same phenotype. However, the same mutations have not yet been carried out in other... 

Mulder EA, et al. Measurement of slow (micros-ms) time scale dynamics in protein side chains by (15)N relaxation dispersion NMR spectroscopy application to Asn and Gin residues in a cavity mutant of T4 lysozyme. J. Am. Chem. Soc. 2001 123 967-975. 

The incorporation of unprotected side-chain derivatives of Asn and Gin derivatives has also been investigated using HATU. In the SPPS of AGP no cyano derivatives of Asn and Gin were detected, however, the incorporation of Asn was not complete, possibly due to the low solubility of Fmoc-Asn-OH in DMF.P l Due to this low solubility of Fmoc-Asn-OH and the unfavorable effects of unprotected side chains on the aggregation behavior of the growing peptide chain, the use of side-chain protection is reconunended (Section 2.3.3).f l For reasons still not clear, even side-chain protection of Asn does not fully eliminate deficiencies in phosphonium- or uronium-mediated couplings. Thus, the incorporation of Fmoc-Asn(Trt)-OH is more satisfactory if one equivalent of HOAt is present during HATU-mediated couplings.P ... 

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