Cysteine residue chemical modification

It is not only the activity that can be altered by incorporation of noncoded amino acids. Introduction of structures possessing certain chemical functions leads to the possibility of highly regioselective modification of enzymes. For example, selective enzymatic modification of cystein residues with compounds containing azide groups has led to the preparation of enzymes that could be selectively immobilized using click chemistry methods [99]. 

Chemical modifications like alkylation with (A-ethylmaleimide (NEM) or oxidation with diamide that inhibit the phosphorylation activity of the enzyme did not seem to have any significant effect on the high affinity binding site when the enzyme was solubilized in the detergent decyl-PEG [69,41]. However, in the intact membrane these treatments reduced the affinity by a factor of 2-3. The reduction of the affinity was exclusively due to modification of the cysteine residue at position 384 in the B domain [69]. Apparently, the detergent effects the interaction between the B and C domains. 

Figure 8.5 Chemical transformation of omeprazole to the corresponding sulphenamide under add conditions and the subsequent modification of an enzyme cysteine residue by the sulphenamide.

Cysteine is the most frequently used residue for selective chemical modification of proteins due to its relatively low abundance in proteins and the increased nucleophilicity of the thiol group relative to other natural amino acid side chains. The intrinsic selectivity is low unless no cysteine is present or unless all unwanted cysteines... 

This covalent bonding arises as a result of biochemical oxidation of the thiol groups in two cysteine residues, and it may also be achieved chemically with the use of mild oxidizing agents. This modification of thiol groups may thus loop a polypeptide chain or cross-link two separate chains. It also significantly modifies the properties of a... 

Tonazzi, A., Giangregorio, N., Indiveri, C. and Palmieri, F. 2005. Identification by site-directed mutagenesis and chemical modification of three vicinal cysteine residues in rat mitochondrial camitine/acylcarnitine transporter. Journal of Biological Chemistry, 280 19607-19612. 

Selective chemical change of the serine—OH group to cysteine—SH in enzymes can be performed with extremely reactive serine residues in the active sites by the use of phenylmethylsulfonyl fluoride and, subsequently, thioacetic acid (Polgar and Bender, 1966). This selective chemical modification demonstrates the essential role of an—OH... 

There is, up to now, one exception known to the four kinds of the above mentioned covalent linkages. The prosthetic group of trimethylamine dehydrogenase is linked via the C(6)-atom of the flavin to a cysteinyl residue (Scheme 3, (5)). As mentioned above the less reactivity of C(6) of flavin as compared to that of CH3(8) requires probably some chemical modification of the prosthetic by biologicai means prior to covalent attachment. The C(6)-S-Cysteinyl flavin was synthesized recently starting with 6-nitro flavin which was subsequently reduced to the amino derivative and transformed to the corresponding bromo derivative via diazotation. Reaction of the bromo derivative with cysteine gave the desired 6-S-Cysteinyl derivative... 

The chemical synthesis of the Amanita toxins has presented several problems, in particular those related to the formation of the sulfur bridge. The latter has been explored with model compounds.[2 31 It has been found that the synthesis of the (sulfanyl)indole moiety can be achieved by reacting an indole compound with an alkanesulfenyl chloride. A model tryptathionine compound has been prepared by reacting A-acyl-L-cysteine and /V-acyl-L-tryptophan in the presence of A-chlorosuccinimide in glacial acetic acid at room temperature.[4] The sulfanylation reaction has been subsequently exploited for the selective chemical modification of tryptophan residues in proteins using 2-nitrophenylsulfenyl chlorideJ5 ... 

Flavin-containing mitochondrial MAO-A and MAO-B catalyze the oxidative deamination of neurotransmitters, such as dopamine, serotonin, and norepinephrine in the central nervous system and peripheral tissues. The enzymes share 73% sequence homology and follow the same kinetic and chemical mechanism but have different substrate and inhibitor specificities. Chemical modification experiments provide evidence that a histidine residue is essential for the catalysis. There is also strong evidence that two cysteine residues are present in the active site of MAO. 

The amino acid sequence of the protein, which can be inferred from the published cDNA sequence (Edman et ai, 1985), reveals that each PDI polyjjeptide contains two domains that are closely homologous to thioredoxin. This enzyme has been characterized in detail (Holmgren, 1985) and it has been shown that the cysteine residues in the sequence WCGPCK (residues 31-36) act as the reactive dithiol. Chemical modification studies have shown that the enzyme, like PDI, is inactivated by alkylation at neutral pH, and that only Cys-32 is alkylated (Kallis and... 

In contrast to albumin, hemoglobin and metallothionein, in which gold(l) coordination to cysteine thiol residues has been clearly established by physicochemical methods, the binding of gold to cyclophilin is unusual in that the Et3PAu+ moiety binds to a histidine residue in preference to the two free thiols present in the protein structure. When cysteine-34 of albumin is blocked by chemical modification, EtsPAuCl, but not auranofin, wiU bind to the numerous histidine residues present. 

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