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CDNAs encoding [D-Ala2]deltorphin precursors from skin of Phyllomedusa bicolor also contain genetic information for three dermorphin-related opioid peptides.

ABSTRACT: We present the structure of four precursors for [D-Ala2]deltorphins I and II as deduced from cDNAs cloned from skin of the frog Phyllomedusa bicolor. These contain the genetic information for one copy of [D-Ala2]deltorphin II and zero, one, or three copies of [D-Ala2]deltorphin I. In each case, the D-alanine of the end product is encoded by a normal GCG codon for L-alanine. In addition, the existence of three peptides related to dermorphin was predicted from the amino acid sequence of the precursors. These peptides were synthesized with a D-alanine in position 2 and their pharmacological properties were tested. Two of them, [Lys7]dermorphin-OH and [Trp4,Asn7]dermorphin-OH, were found to have roughly the same affinity and selectivity for mu-type opioid receptors as dermorphin.

SUBMITTER: Richter K

PROVIDER: S-EPMC54213 | biostudies-other | 1990 Jun

REPOSITORIES: biostudies-other

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cDNAs encoding [D-Ala2]deltorphin precursors from skin of Phyllomedusa bicolor also contain genetic information for three dermorphin-related opioid peptides.

Richter K K Egger R R Negri L L Corsi R R Severini C C Kreil G G

Proceedings of the National Academy of Sciences of the United States of America 19900601 12

We present the structure of four precursors for [D-Ala2]deltorphins I and II as deduced from cDNAs cloned from skin of the frog Phyllomedusa bicolor. These contain the genetic information for one copy of [D-Ala2]deltorphin II and zero, one, or three copies of [D-Ala2]deltorphin I. In each case, the D-alanine of the end product is encoded by a normal GCG codon for L-alanine. In addition, the existence of three peptides related to dermorphin was predicted from the amino acid sequence of the precur ...[more]

PMID: 2352951

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Project description:BackgroundSkin photodamage is associated with ultraviolet- (UV-) induced reactive oxygen species (ROS) overproduction and nuclear factor erythroid 2-related factor 2 (Nrf2) inactivation. In our previous study, skin-derived precursors (SKPs) were shown to ameliorate a UV-induced damage in mice, probably through Nrf2 activation and ROS scavenging.ObjectiveTo clarify the mechanism underlying the photoprotective effect of SKPs against UV-induced damage in a three-dimensional (3D) skin model.MethodsThe Nrf2 gene in SKPs was modified using lentiviral infection, and 3D skin models were reconstructed with keratinocytes and fibroblasts on the basis of type I collagen. Subsequently, these models were divided into the following six groups: normal, model, overexpressed, control, silenced, and negative control groups. Prior to irradiation, respective SKPs were injected into the last four groups. Next, all groups except the normal group were exposed to UVA+UVB. Lastly, the pathological and molecular-biological techniques were employed to determine the parameters. Additionally, LY294002, a PI3K inhibitor, was used to investigate the roles of PI3K/Akt and Nrf2/hemeoxygenase-1 (HO-1) in SKP photoprotection.ResultsNormal 3D skin models appeared as milky-white analogs with a clear, well-arranged histological structure. After the skin was exposed to irradiation, it exhibited cell swelling and a disorganized structure and developed nuclear condensation with numerous apoptotic cells. The expressions of cellular protective genes and Nrf2/HO-1/PI3K/Akt proteins remarkably decreased, which were accompanied by increased oxidative stress and decreased antioxidants (P < 0.05). However, these phenomena were reversed by nrf2-overexpressing SKPs. The 3D skin in the overexpressed group showed mild swelling, neatly arranged cells, and few apoptotic cells. Cellular protective genes and Nrf2/HO-1/PI3K/Akt proteins were highly expressed, and the oxidative biomarkers were remarkably ameliorated (P < 0.05). Nevertheless, the expression of these proteins decreased after LY294002 pretreatment regardless of SKP treatment or not. Meanwhile, there were increases in both UV-induced apoptotic cells and ROS level accompanied with SOD and GPX decrease in the presence of LY294002.ConclusionsEvidence from the 3D skin model demonstrates that the protection of SKPs against UV-mediated damage is primarily via the PI3K/Akt-mediated activation of the Nrf2/HO-1 pathway, indicating that SKPs may be a promising candidate for the treatment of photodermatoses.

Project description:Snake venom metalloproteinases (SVMPs) are a superfamily of zinc-dependent proteases and participate in a number of important biological, physiological and pathophysiological processes. In this work, we simultaneously amplified nine cDNAs encoding different classes of metalloproteinases from glands of four different snake species (Agkistrodon contortrix laticinctus, Crotalus atrox, Crotalus viridis viridis and Agkistrodon piscivorus leucostoma) by RT-PCR with a pair of primers. Among the encoded metalloproteinases, two enzymes (AclVMP-I and AplVMP-I), three enzymes (CaVMP-II, CvvVMP-II and AplVMP-II) and four enzymes (AclVMP-III, CaVMP-III, CvvVMP-III and AplVMP-III) with the characteristic motif (HEXXHXXGXXH) of metalloproteinase belong to type P-I, P-II and P-III enzymes, respectively. Disintegrin domains of CaVMP-II and CvvVMP-II from two Crotatus snakes contain RGD-motif whereas AplVMP-II from Agkistrodon snake has KGD-motif. Instead of R/KGD-motif within disintegrin domain of SVMP-II enzyme, CaVMP-III, CvvVMP-III and AplVMP-III enzymes contain SECD-motif, while AclVMP-III has DDCD-motif in their corresponding position of disintegrin-like domains. There are 12 Cys amino acids in cysterin-rich domains of each P-III enzyme. Moreover, a disintegrin precursor (AplDis) with RGD-motif also simultaneously amplified from the glands of A.p. leucostoma while amplifying AplVMP-II and AplVMP-III, which indicated that different types of SVMPs and related genes are present in a single species of snake and share a consensus sequence at the 3' and 5' untranslated regions. RT-PCR result also showed that P-III is highly expressed in Crotalus snakes than in Agkistrodon snakes. Aligning the deduced amino acid sequence of these enzymes with other SVMPs from GenBank database indicated that this is the first report on the isolation of cDNAs encoding P-II and P-III enzymes from C.v. viridis and A.p. leucostoma snakes. The availability of these SVMP sequences directly facilitated further studies of structure characterization and diversified function analysis.

Dataset Information

CDNAs encoding [D-Ala2]deltorphin precursors from skin of Phyllomedusa bicolor also contain genetic information for three dermorphin-related opioid peptides.

Publications

cDNAs encoding [D-Ala2]deltorphin precursors from skin of Phyllomedusa bicolor also contain genetic information for three dermorphin-related opioid peptides.

Similar Datasets

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