Peptides iodination

Site-specific chemical modifications For examples, methyl esterification that adds 14m.u. for each carboxyl group (i.e. side chains of Asp, Glu or C-terminus in the peptide) iodination that adds 126m.u. for each Tyr. 

The catalytic subunit then catalyzes the direct transfer of the 7-phosphate of ATP (visible as small beads at the end of ATP) to its peptide substrate. Catalysis takes place in the cleft between the two domains. Mutual orientation and position of these two lobes can be classified as either closed or open, for a review of the structures and function see e.g. [36]. The presented structure shows a closed conformation. Both the apoenzyme and the binary complex of the porcine C-subunit with di-iodinated inhibitor peptide represent the crystal structure in an open conformation [37] resulting from an overall rotation of the small lobe relative to the large lobe. 

Thiazolidines have been prepared from /3-aminothiols—for example, cysteine—to protect the —SH and — NH groups during syntheses of peptides, including glu-tathione. Thiazolidines are oxidized to symmetrical disulfides with iodine they do not react with thiocyanogen in a neutral solution. 

Lee PHK, Nguyen TM-D, Chung NN, Schiller PW, Chang KJ. Tyrosine-iodination converts the delta opioid peptide antagonist TIPP to an agonist. Eur J Pharmacol 1995 280 211-214. 

TES-32 is the most abundant single protein product secreted by the parasite. It is also heavily labelled by surface iodination of live larvae (Maizels et al., 1984, 1987), and is known by monoclonal antibody reactivity to be expressed in the cuticular matrix of the larval parasite (Page et al, 1992a). TES-32 was cloned by matching peptide sequence derived from gel-purified protein to an expressed sequence tag (EST) dataset of randomly selected clones from a larval cDNA library (Loukas et al., 1999). Because of the high level of expression of TES-32 mRNA, clones encoding this protein were repeatedly sequenced and deposited in the dataset (Tetteh et al., 1999). Full sequence determination showed a major domain with similarity to mammalian C-type (calcium-dependent) lectins (C-TLs), together with shorter N-terminal tracts rich in cysteine and threonine residues. Native TES-32 was then shown to bind to immobilized monosaccharides in a calcium-dependent manner (Loukas et al., 1999). 

Reported applications of SASD involve modification of lipopolysaccharide (LPS) molecules and studying their interaction with albumin and an antibody directed against LPS (Wollenweber and Morrison, 1985), identification of the murine interleukin-3 receptor and an N-formyl peptide receptor (Sorenson et al., 1986), crosslinking of factor V and Va to iodinated peptides... 

Each bead can iodinate up to 500 pg of tyrosine-containing protein or peptide. This translates into an oxidative capacity of about 0.55 pmol per bead. The rate of reaction can be controlled by changing the number of beads that are used and altering the sodium iodide concentration added to the reaction. Reaction volumes of 100-1,000 pi are possible per bead. The following protocol is suggested for iodinating proteins. Optimization should be done to determine the best incorporation level to obtain good radiolabel incorporation with retention of protein activity. 

Add from 5 pg to 500 pg of a tyrosine-containing peptide or protein dissolved in iodination buffer to the reaction mixture. 

Peptides in Human Urine (Skarzynski and Samecka-Keller), 5, 107 Protein Bound Iodine (Chaney), I, 82 Radioactive Iodine-131 in the Diagnosis of Hyperthyroidism, Blood Plasma Levels of (Silver), 1, 111 Transaminase Activities of Serum and Body Fluids, the Clinical Significance of Alterations in (Wroblew-ski), 1, 314... 

FIGURE 5.22 (A) Reaction of an Fmoc-amino acid with 2-chlorotrityl chloride resin.56 The ester bond formed is cleavable by the mild acid, which does not affect tert-butyl-based protectors. (B) Generation of a protected peptide containing cystine by detachment of a chain, deprotection of cysteine residues, and oxidation of the sulfhydryls by the reagent containing iodine. The cations produced are trapped by CF3CH2OH. 

D Sahal. Removal of iodine by solid phase adsorption to charcoal following iodine oxidation of acetamidomethyl-protected peptide precursors to their disulfide bonded products oxytocin and a Pre-S, peptide of hepatitis B illustrate the method. Int J Pept Res 53, 91, 1999. 

FIGURE 6.23 The synthesis of insulin, starting with a cystine-containing peptide. [Kamber et al., 1977]. Moc = methoxycarbonyl, Bpoc = biphenylisopropoxycarbonyl, Trt = trityl, Acm = acetamidomethyl. (a) HOBt-assisted carbodiimide-mediated coupling (b) removal of Trt by HC1 in CF3CH2OH-CH2Cl2 (9 1) at pH 3.5 (c) removal of Bpoc by CF3CH2OH-CH2 Cl2 (9 1) at 60°C (d) removal of Acm and oxidation by iodine. 

Kamber B, Hartmann A, Eisler K, Riniker B, Rink H, Sieber P, Rittel W. The synthesis of cystine peptides by iodine oxidation of 5-trityl-cysteine and S-acetamidomethyl-cysteine peptides. Helv Chim Acta 1980 63 899-915. 

Thyroid epithelial cells synthesize and secrete T4 and T3 and make up the functional units of thyroid glandular tissue, the thyroid follicles. Thyroid follicles are hollow vesicles formed by a single layer of epithelial cells that are filled with colloid. T4,T3, and iodine are stored in the follicular colloid. T4 and T3 are derived from tyrosyl residues of the protein thyroglobulin (Tg). Thyroid follicular cells synthesize and secrete Tg into the follicular lumen. Thyroid follicular cells also remove iodide (I ) from the blood and concentrate it within the follicular lumen. Within the follicles, some of the tyrosyl residues of Tg are iodinated, and a few specific pairs of iodoty-rosyl residues may be coupled to form T4 and T3. Thus, T4, T3, and iodine (in the form of iodinated tyrosyl residues) are found within the peptide structure of the Tg that is stored in the follicular lumen. 

The secretion of T4 and T3 requires the uptake of follicular contents across the follicular cell apical membrane, the enzymatic release of T4 and T3 from peptide linkage within Tg, and the transport of T4 and T3 across the follicular cell basal membrane to the blood. Several of the steps in synthesis and secretion of T4 and T3 may be compromised by iodine deficiency or disease and can be blocked selectively by a variety of chemicals and drugs. 

T4, T3, MIT, and DIT are stored outside the cell in the follicular colloid in peptide linkage within the Tg molecules. In normal humans on an iodine-sufficient diet, Tg makes up approximately 30% of the mass of the thyroid gland and represents a 2- to 3-month supply of hormone. The total amount of iodine contained as T4, T3, MIT, and DIT within Tg varies with the dietary iodine intake. 

The prototype of oxidizing agents that convert S-protected cysteine peptides directly into cystine peptides (Scheme 1, route B) is iodine. This procedure was initially introduced by Kamber and Rittel for the oxidation of 5-Trt cysteine peptides 66 and has since been extended to the 5-Acm/45,67 5-Tacm/68 5-Phacm/69-71 and 5-Tmb derivatives/64 It can also be... 

A reasonable mechanism for the iodine oxidation of 5-Trt cysteine peptides is given in Scheme 6. 45 Reaction of iodine with the divalent sulfur atom leads to the iodosulfonium ion 5 which is then transformed to the sulfenyl iodide 6 and the trityl cation. Sulfenyl iodides are also postulated as intermediates in the iodine oxidation of thiols to disulfides. The disulfide bond is then formed by disproportionation of two sulfenyl iodides or by reaction between the electrophilic sulfur atom of R -S-I and the nucleophilic S-atom of a second R -S-Trt molecule. The proposed mechanism suggests that any sulfur substitution (i.e., thiol protecting group) capable of forming a stabilized species on cleavage, such as the trityl cation, can be oxidatively cleaved by iodine. 

Table 3 The Solvent Effect on lxn Values for the Iodine Oxidation of 5-Trt- and 5-Acm-Protected Peptide Derivatives 451...

The iodine oxidation method has also been applied in one-pot synthetic strategies to mono-disulfide peptides in which the bis-cysteine peptide is cleaved from the resin, depro-tected, and oxidized in a single step.163 64 A comparative analysis of the iodine procedure (TFA/10 equiv I2) with other TFA/oxidant mixtures using oxytocin as the target model peptide clearly revealed formation of side products to a significant extent. 85 ... 

Much of the experience gained in the field of solution-phase oxidative deprotection of cysteine peptides can be applied to solid-phase chemistry. For this purpose the iodine, thallium(III), and chlorosilane/sulfoxide procedures have been usually employed,163 107-1121 generally, the results obtained with the thallium(III) reagent are better than with iodine. 

A general procedure for the iodine reaction in the solid phase is shown in Scheme 9. Both Boc and Fmoc chemistry can be used to assemble the linear S-protected bis-cysteine peptides and for thiol protection the well-established Acm and Trt groups are usually used. Suitable solvents for the thiol-deprotection/oxidation step by iodine to form the disulfide are CH2C12, DMF, or aqueous AcOH. The final deblocking and cleavage from the resin is carried out under standard conditions. Modification at sensitive amino acid residues caused... 

---