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Publication date: Available online 17 January 2014 Source:Peptides

Author(s): Isabelle M.E. Lacroix , Eunice C.Y. Li-Chan

Inhibition of the enzyme dipeptidyl peptidase (DPP)-IV is one of the strategies used for the treatment of type 2 diabetes. In the present study, pepsin-treated whey protein isolate (WPI) and α-lactalbumin displaying DPP-IV inhibitory activity were fractionated by successive chromatographic steps and the resulting active fractions analyzed for their constituent peptides by liquid chromatography–electrospray ionization-tandem mass spectrometry. Among the identified sequences, 24 peptides derived from α-lactalbumin and 11 from β-lactoglobulin were synthesized and their effects on DPP-IV activity assessed. The most potent fragments, LKPTPEGLD and LKPTPEGLDLEIL (IC50 =45 and 57μM, respectively), were found to inhibit DPP-IV in an un-competitive manner. Although several of the peptides tested showed some inhibitory activity, only two were as effective as the un-fractionated WPI hydrolysate and none were as potent as the un-fractionated α-lactalbumin hydrolysate. The peptides’ structural features, including length and amino acid composition, were found to impact their inhibitory activity. This study provides new insights on the active components responsible for the DPP-IV inhibitory activity of pepsin-treated whey proteins.

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Publication date: February 2014 Source:Peptides, Volume 52

Author(s): Talat Waseem , Mark Duxbury , Stanley W. Ashley , Malcolm K. Robinson

Little is known about ghrelin's effects on intestinal epithelial cells even though it is known to be a mitogen for a variety of other cell types. Because ghrelin is released in close proximity to the proliferative compartment of the intestinal tract, we hypothesized that ghrelin may have potent pro-proliferative effect on intestinal epithelial cells as well. To test this hypothesis, we characterized the effects of ghrelin on FHs74Int and Caco-2 intestinal epithelial cell lines in vitro. We found that ghrelin has potent dose dependent proliferative effects in both cell lines through a yet to be characterized G protein coupled growth hormone secretagogue receptor (GHS-R) subtype. Consistent with above findings, cell cycle flowcytometric analyses demonstrated that ghrelin shifts cells from the G1 to S phase and thereby promotes cell cycle progression. Further characterization of subcellular events, suggested that ghrelin mediates its pro-proliferative effect through Adenylate cyclase (AC)-independent epidermal growth factor receptor (EGFR) trans-activation and PI3K-Akt phosphorylation. Both these pathways converge to stimulate MAPK, ERK 1/2 downstream. The role of ghrelin in states where intestinal mucosal injury and rapid mucosal repair occur warrants further investigation.

Publication date: Available online 10 January 2014 Source:Peptides

Author(s): Liping Jiang , Meichun Deng , Zhigui Duan , Xing Tang , Songping Liang

Spider venom contains a very valuable repertoire of natural resources to discover novel components for molecular diversity analyses and therapeutic applications. In this study, HWTX-XI toxins from the spider venom glands of Ornithoctonus huwena which are Kunitz-type toxins (KTTs) and were directly cloned, analyzed and functionally characterized. To date, the HWTX-XI superfamily consists of 38 members deduced from 121 high-quality expressed sequence tags, which is the largest spider KTT superfamily with significant molecular diversity mainly resulted from cDNA tandem repeats as well as focal hypermutation. Among them, HW11c40 and HW11c50 may be intermediate variants between native Kunitz toxins and sub-Kunitz toxins based on evolutionary analyses. In order to elucidate their biological activities, recombinant HW11c4, HW11c24, HW11c27 and HW11c39 were successfully expressed, further purified and functionally characterized. Both HW11c4 and HW11c27 display inhibitory activities against trypsin, chymotrypsin and kallikrein. Moreover, HW11c4 is also an inhibitor relatively specific for Kv1.1 channels. HW11c24 and HW11c39 are found to be inactive on chymotrysin, trypsin, kallikrein, thrombin and ion channels. These findings provide molecular evidence for toxin diversification of the HWTX-XI superfamily and useful molecular templates of serine protease inhibitors and ion channel blockers for the development of potentially clinical applications.

Publication date: February 2014 Source:Peptides, Volume 52

Author(s): Yingchun Jiang , Xinping Xi , Lilin Ge , Nan Yang , Xiaojuan Hou , Jie Ma , Chengbang Ma , Yuxin Wu , Xiaoxiao Guo , Renjie Li , Mei Zhou , Lei Wang , Tianbao Chen , Chris Shaw

While bradykinin has been identified in the skin secretions from several species of amphibian, bradykinin-related peptides (BRPs) are more common constituents. These peptides display a plethora of primary structural variations from the type peptide which include single or multiple amino acid substitutions, N- and/or C-terminal extensions and post-translational modifications such as proline hydroxylation and tyrosine sulfation. Such modified peptides have been reported in species from many families, including Bombinatoridae, Hylidae and Ranidae. The spectrum of these peptides in a given species is thought to be reflective of its predator profile from different vertebrate taxa. Here we report the isolation of BRPs and parallel molecular cloning of their respective biosynthetic precursor-encoding cDNAs from the skin secretions of the Mexican leaf frog (Pachymedusa dacnicolor), the Central American red-eyed leaf frog (Agalychnis callidryas) and the South American orange-legged leaf frog (Phyllomedusa hypochondrialis). Additionally, the eight different BRPs identified were chemically synthesized and screened for bioactivity using four different mammalian smooth muscle preparations and their effects and rank potencies were found to be radically different in these with some acting preferentially through bradykinin B1-type receptors and others through B2-type receptors.

Publication date: Available online 7 January 2014 Source:Peptides

Author(s): Joshua L. Fuqua , Ofelia M. Littrell , Martin Lundblad , Jadwiga Turchan-Cholewo , Lina G. Abdelmoti , Emilia Galperin , Luke H. Bradley , Wayne A. Cass , Don M. Gash , Greg A. Gerhardt

Glial cell-line derived neurotrophic factor (GDNF) has demonstrated robust effects on dopamine (DA) neuron function and survival. A post-translational processing model of the human GDNF proprotein theorizes the formation of smaller, amidated peptide(s) from the proregion that exhibit neurobiological function, including an 11-amino-acid peptide named dopamine neuron stimulating peptide-11 (DNSP-11). A single treatment of DNSP-11 was delivered to the substantia nigra in the rat to investigate effects on DA-neuron function. Four weeks after treatment, potassium (K+) and d-amphetamine evoked DA release were studied in the striatum using microdialysis. There were no significant changes in DA-release after DNSP-11 treatment determined by microdialysis. Dopamine release was further examined in discrete regions of the striatum using high-speed chronoamperometry at 1-, 2-, and 4-weeks after DNSP-11 treatment. Two weeks after DNSP-11 treatment, potassium-evoked DA release was increased in specific subregions of the striatum. However, spontaneous locomotor activity was unchanged by DNSP-11 treatment. In addition, we show that a single treatment of DNSP-11 in the MN9D dopaminergic neuronal cell line results in phosphorylation of ERK1/2, which suggests a novel cellular mechanism responsible for increases in DA function.

Publication date: February 2014 Source:Peptides, Volume 52

Author(s): Yimin Shao , Geheng Yuan , Yan Feng , Junqing Zhang , Xiaohui Guo

Glucagon-like peptide-1 (GLP-1) has been proved to have effects of anti-hyperglycemia and β-cell preservation. However, it is still unclear whether there are differences between early and late GLP-1 intervention in type 2 diabetes mellitus (T2DM). We divided the mice into 5 groups: early treated group (n =7, 8-week old, fasting glucose>10mmol/l), late treated group (n =7, 10-week old, fasting glucose>20mmol/l), early control group (n =7), late control group (n =7) and wild type group (n =7). Treated group was injected with liraglutide (a GLP-1 analog) 300μg/kg bid for 4 weeks, while control group was given saline at the same time. The results showed that compared with control group, food intake and body weight gain were reduced in both early and late treated group (p <0.05), and there was no significance between the two treated groups. Early liraglutide intervention showed better improvements in glucose control, acute insulin response to glucose (AIRg) and disposition index (before vs. after treatment, AIRg 1.01±0.53 vs. 2.98±0.63, disposition index 10.81±0.89 vs. 27.4±2.15) than late intervention (AIRg 0.99±0.02 vs. 1.41±0.32, disposition index 3.47±0.38 vs. 6.43±1.62, p =0.001). The histopathology of the pancreas showed the estimated β-cell mass (BCM) was increased more in early treated group than that in late one (0.03 vs. 0.01g). Expressions of the proliferation related genes PDX-1, MafA and GLP-1 receptor (GLP-1R) in early treated group were 1.81, 2.57 and 1.59 times as much as that in late treated group. In conclusion, early liraglutide intervention was better in glucose control, β-cell function improvement and β-cell mass preservation.

Publication date: Available online 5 January 2014 Source:Peptides

Author(s): R. Elwyn Isaac , Young-Joon Kim , Neil Audsley

The male sex peptide (SP) of Drosophila melanogaster has wide ranging effects on females, including rejection of courting males, increased egg production, changes to the feeding habit, increased synthesis of antimicrobial peptides and elevated locomotor activity during day-time. The peptide activates receptors in sensory neurons of the female reproductive tract and can also traverse into the hemolymph and reach the central nervous system. The SP receptor involved in rejection and egg-laying responses has been shown to be identical to the receptor for the evolutionary conserved myoinhibitory peptides (MIPs) that function as neuropeptides in both males and females. Intriguingly, MIPs cannot substitute for SP when either expressed in the male accessory glands or injected into virgin females. MIPs are linear peptides with an amidated C-terminus which protects them from cleavage by carboxypeptidases, but leaves them exposed to potential attack from aminopeptidase and endopeptidase activities. In contrast, the SP region responsible for eliciting the post-mating response is cyclic and has several hydroxyproline residues N-terminal to the disulfide bridge which is expected to protect the biological activity of SP from peptidases of the male accessory gland and seminal fluid. We now present in vitro data showing that SP is metabolically stable, whereas MIPs are much more susceptible to degradation by peptidases of the male accessory gland and the hemolymph of virgin female D. melanogaster. SP has evolved relatively recently as a MIP receptor ligand that is particularly well adapted to surviving in the hostile degradome of the male accessory gland and seminal fluid.

Publication date: February 2014 Source:Peptides, Volume 52

Author(s): Luhong Xu , Wenhao Qian , Wenhua Li , Jiali Liu , Haiyan He , Gonghao Li , Yan Cao , Yaren Yu

The association between the levels of N-terminal pro-brain natriuretic peptide (NT-proBNP) and the severity of coronary artery disease (CAD) diagnosed by coronary angiography and other approaches has been investigated. The clinical application of NT-proBNP is restricted by the drawbacks of these techniques now available in screening out patients who need intensive or conservative treatment. Fractional flow reserve (FFR) is superior to coronary angiography and other functional indicators. Accordingly, we designed to investigate the association between NT-proBNP and myocardial ischemia from the perspective of anatomy and physiology in patients with unstable angina and preserved left ventricular function. Plasma samples were collected from 110 patients and NT-proBNP levels were measured by radioimmunoassay. The severity of coronary artery stenosis in patients was measured by coronary angiography and FFR. Stenosis ≥50% in the left main artery or stenosis of 70%, and fractional flow reserve (FFR) ≤0.80 in one or more coronary branches with diameter ≥2mm were defined as “positive”, which require revascularization. NT-proBNP levels increased progressively between patients with negative and positive angiographic results (p <0.05), and between FFR-negative and FFR-positive patients (p <0.05). A significant correlation was observed between logNT-proBNP and logGS (GS=Gensini score, p <0.001). NT-proBNP level serves as a predictor of positive results of angiographic stenosis and FFR, with the area under the receiver operating characteristic curve being 0.697 and 0.787, respectively. NT-proBNP levels are correlated with the severity of anatomic coronary obstruction and inducible myocardial ischemia, but NT-proBNP per se is insufficient to identify clinically significant angiographic and physiological stenoses.

Publication date: January 2014 Source:Peptides, Volume 51

Publication date: February 2014 Source:Peptides, Volume 52

Author(s): Melgious Jin Yan Ang , Zhitao Li , Huichang Annie Lim , Fui Mee Ng , Siew Wen Then , John Liang Kuan Wee , Joma Joy , Jeffrey Hill , C.S. Brian Chia

The Murray Valley encephalitis virus (MVEV) and the West Nile virus (WNV) are mosquito-borne single-stranded RNA Flaviviruses responsible for many cases of viral encephalitis and deaths worldwide. The former is endemic in north Australia and Papua New Guinea while the latter has spread to different parts of the world and was responsible for a recent North American outbreak in 2012, resulting in 243 fatalities. There is currently no approved vaccines or drugs against MVEV and WNV viral infections. A plausible drug target is the viral non-structural NS2B/NS3 protease due to its role in viral replication. This trypsin-like serine protease recognizes and cleaves viral polyproteins at the C-terminal end of an arginine residue, opening an avenue for the development of peptide-based antivirals. This communication compares the P2 and P3 residue preferences of the MVEV and WNV NS2B/NS3 proteases using a series of C-terminal agmatine dipeptides. Our results revealed that both viral enzymes were highly specific toward lysines at the P2 and P3 positions, suggesting that a peptidomimetic viral protease inhibitor developed against one virus should also be active against the other.

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