2-iodoacetamide

The anomalous iodoacetamide-fluoride reaction violates this rule, in that a less stable -halonium complex (18) must be involved, which then opens to (19) in the Markownikoff sense. This has been rationalized in the following way estimates of nonbonded destabilizing interactions in the possible products suggest that the actual product (16) is more stable than the alternative 6)5-fluoro-5a-iodo compound, so the reaction may be subject to a measure of thermodynamic control in the final attack of fluoride ion on the iodonium intermediate. To permit this, the a- and -iodonium complexes would have to exist in equilibrium with the original olefin, product formation being determined by a relatively high rate of attack upon the minor proportion of the less stable )9-iodonium ion. 

Draw a simple mechanism for the reaction of a cysteine sulfhydryl group with iodoacetamide. 

A-Acetylmetbionine Derivative(Chart 9). Cleaved by alkylation of the thioether with iodoacetamide followed by cyclization. 

The reaction of iodoacetamide and its N-substituted derivatives with the Ca -ATPase... 

Cysteine Iodoacetamide, maleimides, Ellman s reagent, p-hydroxymercuribenzoate... 

A failure by one of us to take fully into account the presence of inactivated xanthine oxidase, leading to misinterpretation of incomplete reaction of enzyme with iodoacetamide and hence to the apparently erroneous conclusion, that the two FAD molecules in the enzyme were non-equivalent (72), may serve as a warning to others. This reagent has since been shown to alkylate the flavin of reduced xanthine oxidase molecules, whether these are of the active or inactivated forms (73). Thus, under conditions where little of the inactivated form is reduced, the reagent becomes a specific one for the active enzyme (20). In the original experiments (59, 72) the content of active enzyme was, by coincidence, rather close to half of the total enzyme present. Thus, the presence of inactivated enzyme, rather than a lack of reactivity of one... 

A thiol-reactive squaraine 36 (iodoacetamide) that displays fluorescence emission above 650 nm was used to develop a reagentless glucose monitoring assay [102]. 

When complex 29 is treated with Nal in acetone solution, the corresponding iodoacetamide complex 30 is formed in situ. This complex reacts with sulfhydryl groups to form the rhena-labeled products 31 [Eq. (6)]. 

Thus, iodoacetamide has the highest reactivity toward cysteine sulfhydryl residues and may be directed specifically for —SH blocking. If iodoacetamide is present in limiting quantities (relative to the number of sulfhydryl groups present) and at slightly alkaline pH, cysteine modification will be the exclusive reaction. For additional information on a-haloacetate reactivities and a protocol for blocking, see Section 4.2 (this chapter). 

Figure 5.33 Benzophenone-4-iodoacetamide reacts with sulfhydryl-containing compounds to give thioether linkages. Subsequent photoactivation of the benzophenone residue gives a highly reactive triplet-state ketone intermediate. The energized electron can insert in active C—H or N—H bonds to give covalent crosslinks.

Benzophenone-4-iodoacetamide is water-insoluble and should be pre-dissolved in DMF or another organic solvent prior to adding an aliquot to an aqueous reaction mixture. Stock solutions may be prepared and stored successfully if protected from light. 

Benzophenone-4-iodoacetamide has been used to study the 100-KDa U5 snRNP protein (hPrp28p) and its interactions (Ismaili et al., 2001). 

The spectral properties of these derivatives are similar to native rhodamine. The excitation maximum occurs at about 543 nm and its emission peak at 567nm, producing light in the orange-red region of the spectrum. The extinction coefficient of tetramethylrhodamine-5-(and-6)-iodoacetamide in methanol at its wavelength of maximum absorptivity, 542 nm, is 81,000M-1cm-1. 

Figure 9.18 This iodoacetamide derivative of tetramethylrhodamine can be used to label sulfhydryl groups via thioether bond formation.

Prepare a 20mM tetramethylrhodamine-5-(and-6)-iodoacetamide solution by dissolving 11.3 mg/ml of DMF. Prepare fresh and protect from light. 

DCIA is 7-diethylamino-3-[(4 -(iodoacetyl)amino)phenyl]-4-methylcoumarin, a derivative of the basic aminomethylcoumarin structure that contains a sulfhydryl-reactive iodoacetyl group and a diethyl substitution on its amine. This particular coumarin derivative is among the most fluorescent UV-excitable iodoacetamide probes available (Sippel, 1981) (Invitrogen). 

Figure 9.25 DCIA can modify sulfhydryl groups through its iodoacetamide group to form thioether linkages.

Figure 9.34 The long side chain of this BODIPY derivative contains a sulfhydryl-reactive iodoacetamide group that can couple to a thiol group to form a thioether bond.

Figure 9.35 The iodoacetamide group of this BODIPY fluorophore can react with sulfhydryl-containing molecules to form thioether linkages.

One Lucifer Yellow derivative is available for labeling sulfhydryl-containing molecules. Lucifer Yellow iodoacetamide is a 4-ethyliodoacetamide derivative of the basic disulfonate aminonaph-thalimide fluorophore structure (Invitrogen). The iodoacetyl groups react with —SH groups in proteins and other molecules to form stable thioether linkages (Figure 9.42). 

Figure 9.42 Lucifer Yellow iodoacetamide can be used to label sulfhydryl-containing molecules, forming thioether bonds.

The spectral characteristics of Lucifer Yellow iodoacetamide produce luminescence at somewhat higher wavelengths than the green luminescence of fluorescein, thus the yellow designation in its name. The excitation maximum for the probe occurs at 426 nm and its emission at 530 nm. The rather large Stoke s shift makes sensitive measurements of emission intensity possible without interference by scattered excitation light. The 2-mercaptoethanol derivative of the fluorophore has an extinction coefficient at pH 7 of about 13,000 M cm-1 at 426nm. 

Figure 9.54 The iodoacetamide derivative of DPA has been used to create a chelating polymer of lanthanide metals using poly-L-lysine as the backbone.

Figure 11.10 This biotinylation reagent reacts with sulfhydryl groups through its iodoacetamide end to form thioether bonds.

---