A-iodoacetamide

In another experiment (J ) we treated ozone-resistant and ozone-susceptible varieties of tobacco with toxic doses of oc-iodoacetic acid, and a-iodoacetamide, both sulfhydryl-binding reagents. The symptoms produced by both compounds were similar to those produced by ozone. The severity of the injury also paralleled ozone resistance (Table II). The degree of injury caused by these two compounds also paralleled the ozone susceptibility of leaves of different ages on the same plant. The uppermost, youngest, leaves appear to be most resistant to both the sulfhydryl-binding reagents and to ozone. 

Over a decade ago, work on the enzyme aldolase reductase elegantly demonstrated this point. The noncovalent inhibitor alrestatin was modified to contain various electrophiles a-chloroacetamide, a-bromoacetamide or a-iodoacetamide. Noncovalent interactions between inhibitors and protein would not have changed, but molecules behaved differently based on the electrophile the weakest showed reversible inhibition, whereas the iodoacetamide displayed almost complete irreversible inhibition.1401 These results are an important warning if a reaction is too facile, irreversible reactions can obscure true binding affinities. 

Figure S. Modifications of thioether groups (a) hydrogen peroxide (b) photooxidation (c) performic acid (a) iodoacetamide.

SYNS a-IODOACETAMIDE 2-IODOACETAMIDE MONOIODOACETAMIDE SURAUTO USAF D-1... 

The EtsB-induced halogen atom transfer radical cyclization reaction is a successful application. Cyclization of iodo acetal 44 afforded the tetrahydrofuran derivative 45 in almost quantitative yield (Scheme 26) [32]. Et3B also induced radical cyclization of A -allylic a-iodoacetamide to give y9-iodomethyl-y-lactam via an atom transfer process [34]. The reaction of 46 prepared from 2-prolinol proceeded smoothly within 10 min in boiling benzene in the presence of Et3B to yield IR, 8S)-... 

The completion of the synthesis of aspidospeimidine relied on a protocol introduced by Heathcock [72] for annulating the E-ring to the ABCD-framework of 228. Thus, as shown in Scheme 23, acylation of target compound 242 with a-chloroacetyl chloride followed by a Finkelstein reaction afforded the a-iodoacetamide 243 that upon treatment with silver triflate effected an intramolecular alkylation reaction and the formation of the isoindole-based lactam 244. Finally, reduction of this last compound with hthium aluminium hydride gave the racemic modification of aspidospermidine (228) (32%). 

Fig. 8.5. Densitometer trace of electrophoretic separation of species trapped by (A) iodoacetamide, (B) iodoacetate (courtesy of Creighton, 1974b). Reduced and native form of BPTI are R and N respectively.

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. 

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]. 

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.

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 9.34 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.

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

To block unreacted sulfhydryl groups, add iodoacetamide to the solution to a final concentration of 2 mM. 

Figure 22.30 An iodoacetamide derivative of PE containing an extended spacer arm can be constructed through a carbodiimide coupling of iodoacetic acid to PE, followed by reaction with 2-mercaptoethylamine, and finally another reaction with iodoacetate.

Figure 25.18 An amine-derivative of dextran may be coupled with iodoacetic acid using a carbodiimide reaction to produce a sulfhydryl-reactive iodoacetamide polymer.

Reduce protein disulfides by adding dithiothreitol (DTT) to a final concentration of 5mM and incubate for 30 minutes at 60°C. Add iodoacetamide to a final concentration of 25 mM to alkylate the thiols. React for 1 hour in the dark. 

N. A. Zorin, B. Dimon, J. Gagnon, J. Gaillard, P. Carrier, P. M. Vignais (1996) Inhibition by iodoacetamide and acetylene of the H-D exchange reaction catalyzed by Thiocapsa roseopersicina hydrogenase. Eur. J. Biochem., 241 675-681... 

Abbreviations lAA, iodoacetamide BFA, brefeldin A PMA, phorbol myristate acetate DNP, dinitrophenol ATPase, adinosine triphosphatase GTPase, guanosin triphosphatase. 

FIGURE 4 Effect of sample preparation on the fragmentation of an rMAb observed in (A) SDS-PAGE and (B) CE-SDS with LIF detection. SDS-PAGE lanes (Lane I) molecular weight standards bovine serum albumin at (Lane 2) 8 ng and (Lane 3) 2 ng (Lane 4) rMAb control after alkylation with (Lane 5) iodoacetic acid and (Lane 6) iodoacetamide. (See color plate 4.)... 

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