Gábor K. Tóth

Application of alicyclic β-amino acids in peptide chemistry

The self-organizing β-peptides have attracted considerable interest in the fields of foldamer chemistry and biochemistry. These compounds exhibit various stable secondary structure motifs that can be exploited to construct biologically active substances and nanostructured tertiary structures. The secondary structures can be controlled via the β-amino acid sequence, and cyclic β-amino acid residues play a crucial role in the design. The most important procedures for the preparation of cyclic β-amino acid monomers and peptides are discussed in this tutorial review. Besides the secondary structure design principles, the methods of folded structure detection are surveyed.

Further analysis of the effects of cholecystokinin octapeptides on avoidance behaviour in rats

Experiments were performed to examine the acute effects of cholecystokinin octapeptides and fragments on the active and passive avoidance behaviour of rats following peripheral and central administration. Both the sulphated (CCK-8-SE) and non-sulphated cholecystokinin octapeptide (CCK-8-NS) and also the COOH-terminal tetra-, penta-, hexa- and heptapeptides of cholecystokinin octapeptide facilitated the extinction of active avoidance behaviour and retention of passive avoidance behaviour. This latter effect of cholecystokinin octapeptides was reversed by anxiolytic chlordiazepoxide pretreatment, showing that in these test situations cholecystokinin octapeptides are able to modify fear-motivation or arousal of the animals; their effect is at least partly similar to that of the neuroleptic substance haloperidol. Subcutaneous treatment with CCK-8-SE or CCK-8-NS appeared to be 3-10 times more effective than intraperitoneal treatment. Following intracerebroventricular administration, 100-300 times lower doses were needed to cause a behavioural effect similar to that after subcutaneous injection. Microinjection of CCK-8-SE or CCK-8-NS in the fmol dose range into the nucleus accumbens facilitated the extinction of active avoidance behaviour and attenuated the retention of passive avoidance behaviour, while microinjection of these peptides into the central amygdaloid nucleus caused opposite effects on these behavioural tests. It seems that the neuroleptic-like effects of cholecystokinin octapeptides are mediated through the nucleus accumbens, and the opposite action (non neuroleptic-like) through the central amygdaloid nucleus.

Role of flanking sequences and phosphorylation in the recognition of the simian-virus-40 large T-antigen nuclear localization sequences by importin-a

The nuclear import of simian-virus-40 large T-antigen (tumour antigen) is enhanced via phosphorylation by the protein kinase CK2 at Ser112 in the vicinity of the NLS (nuclear localization sequence). To determine the structural basis of the effect of the sequences flanking the basic cluster KKKRK, and the effect of phosphorylation on the recognition of the NLS by the nuclear import factor importin-alpha (Impalpha), we co-crystallized non-autoinhibited Impalpha with peptides corresponding to the phosphorylated and non-phosphorylated forms of the NLS, and determined the crystal structures of the complexes. The structures show that the amino acids N-terminally flanking the basic cluster make specific contacts with the receptor that are distinct from the interactions between bipartite NLSs and Impalpha. We confirm the important role of flanking sequences using binding assays. Unexpectedly, the regions of the peptides containing the phosphorylation site do not make specific contacts with the receptor. Binding assays confirm that phosphorylation does not increase the affinity of the T-antigen NLS to Impalpha. We conclude that the sequences flanking the basic clusters in NLSs play a crucial role in nuclear import by modulating the recognition of the NLS by Impalpha, whereas phosphorylation of the T-antigen enhances nuclear import by a mechanism that does not involve a direct interaction of the phosphorylated residue with Impalpha.

Design of Peptidic Foldamer Helices: A Stereochemical Patterning Approach

Assembly language: The programmed sequences of stereochemical building blocks lead to novel biomimetic helices. The rational design approach offers new possibilities for creating periodic secondary structures.

Cell-penetrating peptide exploited syndecans.

Cell-penetrating peptides (CPPs) are short peptides capable of translocating across the plasma membrane of live cells and transporting conjugated compounds intracellularly. Fifteen years after discovering the first model cationic CPPs, penetratin and TAT, CPP internalization is still challenging many questions. Particularly it has been unknown whether CPPs enter the cells with or without mediation of a specific surface receptor. Here we report that syndecan-4, the universally expressed isoform of the syndecan family of transmembrane proteoglycans, binds and mediates transport of the three most frequently utilized cationic CPPs (penetratin, octaarginine and TAT) into the cells. Quantitative uptake studies and mutational analyses demonstrate that attachment of the cationic CPPs is mediated by specific interactions between the heparan sulfate chains of syndecan-4 and the CPPs. Protein kinase C alpha is also heavily involved in the uptake mechanism. The collected data give the first direct evidence on the receptor-mediated uptake of cationic CPPs and may replace the long-thought, but already contradicted membrane penetration hypothesis. Thus our study might give an answer for a decade long debate and foster the development of rationalized, syndecan-4 targeted novel delivery technologies.

Decreased serum and red blood cell kynurenic acid levels in Alzheimer's disease

Kynurenine aminotransferases (KAT I and KAT II) are responsible for the transamination of kynurenine (KYN) to form kynurenic acid (KYNA), an excitatory amino acid receptor antagonist. Since these members of the kynurenine pathway (KP) are proposed to be involved in the pathogenesis of Alzheimers dementia (AD), the activities of these enzymes and the levels of these metabolites were measured in the plasma and red blood cells (RBCs) of AD and control subjects together with the inheritance of the apolipoprotein (APOE) epsilon4 allele. KYNA levels were significantly decreased both in the plasma and in the RBCs in AD, but the levels of KYN and the activities of KAT I and KAT II remained unchanged. No association has been found with the possession of the epsilon4 allele. These findings indicate an altered peripheral KP in AD regardless of the APOE status of the probands.

Pituitary adenylate cyclase activating polypeptide protects cardiomyocytes against oxidative stress-induced apoptosis

Pituitary adenylate cyclase activating polypeptide (PACAP) has well-known neuroprotective effects, and one of the main factors leading to neuroprotection seems to be its anti-apoptotic effects. The peptide and its receptors are present also in the heart, but whether PACAP can be protective in cardiomyocytes, is not known. Therefore, the aim of the present study was to investigate the effects of PACAP on oxidative stress-induced apoptosis in cardiomyocytes. Our results show that PACAP increased cell viability by attenuating H2O2-induced apoptosis in a cardiac myocyte culture. PACAP also decreased caspase-3 activity and increased the expression of the anti-apoptotic markers Bcl-2 and phospho-Bad. These effects of PACAP were counteracted by the PACAP antagonist PACAP6-38. In summary, our results show that PACAP is able to attenuate oxidative stress-induced cardiomyocyte apoptosis.

The neuroprotective effects of PACAP in monosodium glutamate-induced retinal lesion involve inhibition of proapoptotic signaling pathways

Pituitary adenylate cyclase activating polypeptide (PACAP) and its receptors are present in the retina and exert several distinct functions. PACAP has well-known neuroprotective effects in neuronal cultures in vitro and against different insults in vivo. Recently we have shown that PACAP is neuroprotective against monosodium glutamate (MSG)-induced retinal degeneration. In the present study we investigated the possible signal transduction pathways involved in the protective effect of intravitreal PACAP administration against apoptotic retinal degeneration induced by neonatal MSG treatment. MSG induced activation of proapoptotic signaling proteins and reduced the levels of antiapoptotic molecules in neonatal retinas. Co-treatment with PACAP attenuated the MSG-induced activation of caspase-3 and JNK, inhibited the MSG-induced cytosolic translocation of apoptosis inducing factor (AIF) and cytochrome c, and increased the level of phospho-Bad. Furthermore, PACAP treatment alone decreased cytosolic AIF and cytochrome c levels, while PACAP6-38 increased cytochrome c release, caspase-3 and JNK activity and decreased phospho-Bad activity. In summary, our results show that PACAP treatment attenuated the MSG-induced changes in apoptotic signaling molecules in vivo and suggest that also endogenously present PACAP has neuroprotective effects. These results may have further clinical implications in reducing glutamate-induced excitotoxicity in several ophthalmic diseases.

Genetic Polymorphisms of NOD1 and IL-8, but not Polymorphisms of TLR4 Genes, Are Associated with Helicobacter pylori-Induced Duodenal Ulcer and Gastritis

Background:  Intracellular pathogen receptor NOD1 is involved in the epithelial cell sensing Helicobacter pylori, which results in a considerable interleukin (IL)‐8 production. The aim of this study was to evaluate the relationship between NOD1 and IL‐8 genetic polymorphisms and the development of H. pylori‐induced gastritis and duodenal ulcer (DU), as compared with TLR4 polymorphisms.

Building β-Peptide H10/12 Foldamer Helices with Six-Membered Cyclic Side-Chains: Fine-Tuning of Folding and Self-Assembly

The ability of the β-peptidic H10/12 helix to tolerate side-chains containing six-membered alicyclic rings was studied. cis-2-Aminocyclohex-3-ene carboxylic acid (cis-ACHEC) residues afforded H10/12 helix formation with alternating backbone configuration. Conformational polymorphism was observed for the alternating cis-ACHC hexamer, where chemical exchange takes place between the major left-handed H10/12 helix and a minor folded conformation. The hydrophobically driven self-assembly was achieved for the cis-ACHC-containing helix which was observed as vesicles ~100 nm in diameter.