Akira Shigenaga

N-->S acyl-transfer-mediated synthesis of peptide thioesters using anilide derivatives.

N-->S acyl-transfer-mediated synthesis of peptide thioesters utilizing an N-aminoacyl-N-sulfanylethylaminobenzoic acid derivative has been examined. The developed synthetic methodology for peptide thioesters is compatible with Fmoc solid-phase peptide synthesis (SPPS).

N-Sulfanylethylanilide Peptide as a Crypto-Thioester Peptide

Native chemical ligation (NCL) has shown great utility in protein chemistry and has yielded impressive success in the preparation of a wide variety of proteins. This methodology requires peptide thioesters that serve as chemoselective acylating agents for N-terminal cysteinyl peptides to afford ligated peptides through a sequence of reactions consisting of S–S and S–N acyl transfers. The susceptibility of the thioester moiety to basic reagents has necessitated the preparation of the key intermediate by Boc-based solid-phase peptide synthesis (Boc-SPPS) without requiring a nucleophile-mediated deprotection procedure. However, the preferred use of Fmoc-based SPPS with piperidine treatment demands the development of a synthetic methodology using peptide thioesters that are compatible with Fmoc chemistry. In this context, many research groups, including ours, have explored an Fmoc-based synthetic protocol for thioesters. Among the reported studies, N–S acyl-transfer-mediated procedures have great potential in Fmoc chemistry. We have also developed an N-sulfanylethylaniline linker that can be used for the acyl-transfer-mediated synthesis of peptide thioesters. Standard Fmoc-SPPS on the sulfanylethylaniline linker followed by N–S acyl transfer under acidic conditions (4 m HCl in DMF) efficiently yielded peptide thioesters (Scheme 1). On the basis of these experimental results, we attempted to utilize an N-terminal cysteinyl N-sulfanylethylanilide (SEAlide) peptide as the middle fragment(s) for sequential NCL, which features the use of more than one thioester fragment. Here, involvement of the SEAlide peptide in the first NCL with a peptide thioester would seem to selectively afford the corresponding ligated SEAlide peptide, which can be used in the second NCL step after conversion of the anilide moiety to the thioester under acidic conditions. The first NCL doubtlessly proceeded; however, contrary to our expectations, a not insignificant amount of cyclic material resulting from the unanticipated intramolecular NCL of the cysteinyl SEAlide peptide was observed (Scheme 2).

Dual Kinetically Controlled Native Chemical Ligation Using a Combination of Sulfanylproline and Sulfanylethylanilide Peptide

Dual kinetically controlled native chemical ligation using a newly developed sulfanylproline-mediated reaction in combination with an N-sulfanylethylanilide peptide was successfully applied to a previously unreported sequential coupling of peptide fragments added simultaneously to the reaction.

Synthesis of a stimulus-responsive processing device and its application to a nucleocytoplasmic shuttle Peptide.

Stimulus-responsive processing (peptide bond cleavage) devices were developed. The processing reaction was triggered by stimulus-induced removal of a PG and the processing products were obtained in good purity. A photo-responsive processing device was successfully applied to develop a nucleocytoplasmic shuttle peptide. (F: fluorophore, NES: nuclear export signal. NLS: nuclear localization signal. PG: stimulusresponsive protective group)

Chemical Synthesis of Biologically Active Monoglycosylated GM2‐Activator Protein Analogue Using N‐Sulfanylethylanilide Peptide

Going to SEA(lide): Total chemical synthesis of a 162-residue glycoprotein analogue of the monoglycosylated human GM2-activator protein (GM2AP) was achieved. Key steps were the use of N-sulfanylethylanilide (SEAlide) peptides in the kinetic chemical ligation synthesis of a large peptide fragment, and a convergent native chemical ligation for final fragment assembly.

CXCL14 is a natural inhibitor of the CXCL12–CXCR4 signaling axis

CXCR4 physically interacts with CXCL14 anti bait coimmunoprecipitation by (View interaction).

Proline cis/trans-Isomerase Pin1 Regulates Peroxisome Proliferator-activated Receptor γ Activity through the Direct Binding to the Activation Function-1 Domain

The important roles of a nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) are widely accepted in various biological processes as well as metabolic diseases. Despite the worldwide quest for pharmaceutical manipulation of PPARγ activity through the ligand-binding domain, very little information about the activation mechanism of the N-terminal activation function-1 (AF-1) domain. Here, we demonstrate the molecular and structural basis of the phosphorylation-dependent regulation of PPARγ activity by a peptidyl-prolyl isomerase, Pin1. Pin1 interacts with the phosphorylated AF-1 domain, thereby inhibiting the polyubiquitination of PPARγ. The interaction and inhibition are dependent upon the WW domain of Pin1 but are independent of peptidyl-prolyl cis/trans-isomerase activity. Gene knockdown experiments revealed that Pin1 inhibits the PPARγ-dependent gene expression in THP-1 macrophage-like cells. Thus, our results suggest that Pin1 regulates macrophage function through the direct binding to the phosphorylated AF-1 domain of PPARγ.

One-Pot/Sequential Native Chemical Ligation Using N-Sulfanylethylanilide Peptide

N-Sulfanylethylanilide (SEAlide) peptides were developed with the aim of achieving facile synthesis of peptide thioesters by 9-fluorenylmethyloxycarbonyl (Fmoc)-based solid-phase peptide synthesis (Fmoc SPPS). Initially, SEAlide peptides were found to be converted to the corresponding peptide thioesters under acidic conditions. However, the SEAlide moiety was proved to function as a thioester in the presence of phosphate salts and to participate in native chemical ligation (NCL) with N-terminal cysteinyl peptides, and this has served as a powerful protein synthesis methodology. The reactivity of a SEAlide peptide (anilide vs. thioester) can be easily tuned with or without the use of phosphate salts. This interesting property of SEAlide peptides allows sequential three-fragment or unprecedented four-fragment ligation for efficient one-pot peptide/protein synthesis. Furthermore, dual-kinetically controlled ligation, which enables three peptide fragments simultaneously present in the reaction to be ligated in the correct order, was first achieved using a SEAlide peptide. Beyond our initial expectations, SEAlide peptides have served in protein chemistry fields as very useful crypto-peptide thioesters.

Synthesis of Amide-Type Fluoroalkene Dipeptide Isosteres by an Intramolecular Redox Reaction

We previously achieved NHC-mediated preparation of ester-type fluoroalkene dipeptide isosteres (ES-FADIs, 4) by an intramolecular redox reaction. In the present study, a cyanide ion-mediated reaction was successfully applied to the conversions of gamma,gamma-difluoro-alpha,beta-enoylsilane 1 or 2 to amide-type fluoroalkene isosteres (AM-FADIs, 5 or 6). The use of catalytic cyanide ion allowed synthesis of chiral auxiliary incorporated FADI 15b which was then subjected to a diastereoselective alpha-alkylation reaction to yield alpha-substituted FADIs 17. Furthermore, the presented amidation protocol was used for straightforward incorporation of FADI into peptidyl resin.

The extreme N-terminal region of human apolipoprotein A-I has a strong propensity to form amyloid fibrils

The N‐terminal 1–83 residues of apolipoprotein A‐I (apoA‐I) have a strong propensity to form amyloid fibrils, in which the 46–59 segment was reported to aggregate to form amyloid‐like fibrils. In this study, we demonstrated that a fragment peptide comprising the extreme N‐terminal 1–43 residues strongly forms amyloid fibrils with a transition to β‐sheet‐rich structure, and that the G26R point mutation enhances the fibril formation of this segment. Our results suggest that in addition to the 46–59 segment, the extreme N‐terminal region plays a crucial role in the development of amyloid fibrils by the N‐terminal fragment of amyloidogenic apoA‐I variants.