Insulin Hot-Spot Analogs Formed with N-Methylated Amino Acid Residues Inhibit Aggregation of Native Hormone

Swiontek, Monika; Waśko, Joanna; Frączyk, Justyna; Gałęcki, Krystian; Kamiński, Zbigniew J.; Kolesińska, Beata

doi:10.3390/molecules24203706

Cited by 9 publications

(7 citation statements)

References 39 publications

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“…On the CD spectra of the HSA-insulin complex ( Figure 9 ), a regular increase was observed in signal intensity at 194 nm and a stronger minimum at 210 and 224 nm. This means that, in the presence, of the native form of insulin does not tend to form amyloid fibrils, for which the β-sheet structure is the main growth factor [ 60 , 67 ]. We can suppose that possible interactions between HSA and the native hormone inhibit undesirable insulin aggregation and increase the content of the α-helix structure in the sample.…”

Section: Resultsmentioning

confidence: 99%

Human Serum Albumin Binds Native Insulin and Aggregable Insulin Fragments and Inhibits Their Aggregation

et al. 2020

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The purpose of this study was to investigate whether Human Serum Albumin (HSA) can bind native human insulin and its A13–A19 and B12–B17 fragments, which are responsible for the aggregation of the whole hormone. To label the hormone and both hot spots, so that their binding positions within the HSA could be identified, 4-(1-pyrenyl)butyric acid was used as a fluorophore. Triazine coupling reagent was used to attach the 4-(1-pyrenyl)butyric acid to the N-terminus of the peptides. When attached to the peptides, the fluorophore showed extended fluorescence lifetimes in the excited state in the presence of HSA, compared to the samples in buffer solution. We also analyzed the interactions of unlabeled native insulin and its hot spots with HSA, using circular dichroism (CD), the microscale thermophoresis technique (MST), and three independent methods recommended for aggregating peptides. The CD spectra indicated increased amounts of the α-helical secondary structure in all analyzed samples after incubation. Moreover, for each of the two unlabeled hot spots, it was possible to determine the dissociation constant in the presence of HSA, as 14.4 µM (A13–A19) and 246 nM (B12–B17). Congo Red, Thioflavin T, and microscopy assays revealed significant differences between typical amyloids formed by the native hormone or its hot-spots and the secondary structures formed by the complexes of HSA with insulin and A13–A19 and B12–B17 fragments. All results show that the tested peptide-probe conjugates and their unlabeled analogues interact with HSA, which inhibits their aggregation.

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Section: Resultsmentioning

confidence: 99%

Human Serum Albumin Binds Native Insulin and Aggregable Insulin Fragments and Inhibits Their Aggregation

et al. 2020

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“…As expected, in many cases the incorporation of one or two N-methylated amino acid residues into the amylin fragments completely eliminated or reduced their susceptibility to form amyloids. [37] For fragment 23 -27, it was necessary to incorporate two N-methylated amino acid residues into the peptide sequence to eliminate its susceptibility to aggregation. This is consistent with the data presented by Kapurniotu.…”

Section: Resultsmentioning

confidence: 99%

“…The procedures described in the previous works [31,37] were used for the syntheses of peptides 1-9. All synthetic procedures for synthesis of HÀ H(NÀ Me) SSNNÀ OH (1), HÀ HS(N-Me)SNNÀ OH (2), HÀ H(NÀ Me) S(NÀ Me)SNNÀ OH (3), HÀ F(NÀ Me)GAILÀ OH (4), HÀ FGA (NÀ Me)ILÀ OH (5) and HÀ F(NÀ Me)GA(NÀ Me)ILÀ OH (6), HÀ G(NÀ Me)SNTYÀ NH 2 (7), HÀ GSNT(NÀ Me)YÀ NH 2 (8), HÀ G(NÀ Me)SNT(NÀ Me)YÀ NH 2 (9) are presented in the Supporting Information.…”

Section: Peptide Synthesismentioning

confidence: 99%

N‐Methylated Analogs of hIAPP Fragments 18–22, 23–27, 33–37 Inhibit Aggregation of the Amyloidogenic Core of the Hormone

Rożniakowski

Gałęcki

Wietrzyk

et al. 2020

Chemistry & Biodiversity

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Amylin (hIAPP) aggregation leads to the formation of insoluble deposits and is one of the factors in the development of type II diabetes. The aim of this research was to find N-methylated analogs of the aggregating amylin fragments 18-22, 23-27, and 33-37, which would not themselves be susceptible to aggregation and would inhibit the aggregation of the amyloidogenic cores of the hormone. None of the analogs of fragment 18-22 containing one or two N-methylated amino acid residues showed any tendency to aggregate. Only the peptide HÀ F(NÀ Me)GA(NÀ Me) ILÀ OH (6) derived from the 23-27 hIAPP hot spot did not form fibrous structures. All analogs of the 33-37 amylin fragment were characterized by the ability to form aggregates, despite the presence of N-methylated amino acids in their structures. N-Methylated peptides 1-5 demonstrated inhibitory properties against the aggregation of fragment 18-22. Aggregation of the amyloidogenic core of 23-27 was significantly inhibited by N-methylated peptides 1-3 derived from the (18-22) HÀ HSSNNÀ OH fragment and by the HÀ F(N-Me)GA(NÀ Me)ILÀ OH (6) fragment derived from the 23-27 amylin hot spot. Fragment (33-37) HÀ GSNTYÀ NH 2 was found to be inhibited in the presence of N-methylated peptides 1-3 derived from the 18-22 fragment and by the double methylated peptide HÀ F(NÀ Me)GA(NÀ Me)ILÀ OH (6). Research on the possibility of using N-methylated analogs of amyloidogenic amylin cores as inhibitors of hormone aggregation is ongoing, with a focus on finding the minimum concentration of N-methylated peptides capable of inhibiting the aggregation of hIAPP hot spots.

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“…DMT/NMM/TsO − [ 67 , 68 ] ( 1 ) is an effective, and environmentally friendly coupling reagent used for the synthesis of amides, esters, and peptides in solution or solid phase, as well as under microwave-assisted conditions [ 69 ]. It enables the synthesis of peptides using Z-, Boc-, and Fmoc- protected substrates as well as unnatural amino acids.…”

Section: Introductionmentioning

confidence: 99%

Synthesis of Isothiocyanates Using DMT/NMM/TsO− as a New Desulfurization Reagent

2021

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Thirty-three alkyl and aryl isothiocyanates, as well as isothiocyanate derivatives from esters of coded amino acids and from esters of unnatural amino acids (6-aminocaproic, 4-(aminomethyl)benzoic, and tranexamic acids), were synthesized with satisfactory or very good yields (25–97%). Synthesis was performed in a “one-pot”, two-step procedure, in the presence of organic base (Et3N, DBU or NMM), and carbon disulfide via dithiocarbamates, with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium toluene-4-sulfonate (DMT/NMM/TsO−) as a desulfurization reagent. For the synthesis of aliphatic and aromatic isothiocyanates, reactions were carried out in a microwave reactor, and selected alkyl isothiocyanates were also synthesized in aqueous medium with high yields (72–96%). Isothiocyanate derivatives of L- and D-amino acid methyl esters were synthesized, under conditions without microwave radiation assistance, with low racemization (er 99 > 1), and their absolute configuration was confirmed by circular dichroism. Isothiocyanate derivatives of natural and unnatural amino acids were evaluated for antibacterial activity on E. coli and S. aureus bacterial strains, where the most active was ITC 9e.

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Insulin Hot-Spot Analogs Formed with N-Methylated Amino Acid Residues Inhibit Aggregation of Native Hormone

Cited by 9 publications

References 39 publications

Human Serum Albumin Binds Native Insulin and Aggregable Insulin Fragments and Inhibits Their Aggregation

Human Serum Albumin Binds Native Insulin and Aggregable Insulin Fragments and Inhibits Their Aggregation

N‐Methylated Analogs of hIAPP Fragments 18–22, 23–27, 33–37 Inhibit Aggregation of the Amyloidogenic Core of the Hormone

Synthesis of Isothiocyanates Using DMT/NMM/TsO− as a New Desulfurization Reagent

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