A strategy for isolating rare peptides: isolation and sequencing of a large peptide present in a single neuron of the nematode Ascaris suum

Sithigorngul, Paisarn; Nanda, Jennifer Cho; Stretton, A. O. W.

doi:10.1016/s0196-9781(03)00184-0

Cited by 7 publications

(6 citation statements)

References 15 publications

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“…The CHHlike peptides have a high degree of sequence similarity and range in size from 69-78 amino acids (Macins et al 1999;Sithigorngul et al 2003). The family separates into two major classes (type I and type II) by structural, functional, and phylogenetic distinctions.…”

Section: Introductionmentioning

confidence: 99%

Molecular characterization and cell-specific expression of an ion transport peptide in the tobacco hornworm, Manduca sexta

et al. 2007

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The crustacean hyperglycemic hormone (CHH) peptides regulate diverse physiological processes from reproduction to metabolism and molting in arthropods. In insects, the ion transport peptides (ITP), also members of the CHH family, have only been implicated in ion transport. In this study, we sequenced a nucleotide fragment spanning the conserved A1/A2 region of the putative CHH/ITP gene. This fragment was amplified from larval cDNA of the tobacco hornworm, Manduca sexta and showed a high degree of sequence conservation with the same region from other insects and, to a lesser degree, with that of crustacean species, suggesting the presence of a Manduca-specific CHH/ITP mRNA (MasITP mRNA). CHH-like immunocytochemical analyses with two crustacean antisera (from Carcinus maenas and Cancer pagurus) identified the presence of CHH-like immunoreactivity in nervous tissue of all developmental stages, but not in the gut of M. sexta. Specifically, CHH-like peptides localized to paired type IA(2) neurosecretory cells of the pars lateralis of the brain (projecting ipsilaterallly to the corpora cardiaca-allata complex) and to neurosecretory cells and transverse nerves of the ventral nerve cord in larvae, pupae, and adults. The distribution of the putative MasITP peptide shifted during development in a manner consistent with metamorphic reorganization. A comparison of hemolymph equivalents of CHH detected by enzyme-linked immunosorbent assay with CHH-like immunoreactivity in transverse nerves provided evidence for the release of MasITP from the transverse nerves into the hemolymph at insect ecdysis. These data suggest the presence of an insect ITP in M. sexta and a role for this hormone during ecdysis.

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Section: Introductionmentioning

confidence: 99%

Molecular characterization and cell-specific expression of an ion transport peptide in the tobacco hornworm, Manduca sexta

et al. 2007

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“…Using an immunosuppressed mouse immunized with crude homogenate of dissected nervous tissue, a 11 kD peptide was isolated. The antibody was used to monitor the peptide during the purification processes 18 . This evidence reflects the efficiency of mouse immune system which responds to very minute amounts of antigen in the crude extract.…”

Section: Discussionmentioning

confidence: 99%

“…The membrane was incubated for 8 hr in 1:100 dilution recognized only a single neuron in the nervous system was subsequently used for peptide isolation. This approach yielded a unique peptide of molecular weight of 11,542 Da that was present in only a single neuron 18 . In this study we have used a similar approach, by generating monoclonal antibodies specific to eyestalk neuropeptides of P. monodon using sinus gland section and immunosuppression technique and selecting hybridoma clones by immunohistochemistry on the eyestalk sections.…”

Section: Introductionmentioning

confidence: 99%

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Panchan¹,

Sitigorngul²,

Chaivisuthangkura³

et al. 2005

ScienceAsia

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“…The membrane was washed thoroughly with deionized water and dried at room temperature. Immunoreactive fractions were identified by a reddish-brown staining in which the fraction had been added to the membrane [20-22]. The immunoreactive fractions were further analyzed by MALDI-FTMS.…”

Section: Methodsmentioning

confidence: 99%

Immunoaffinity-based mass spectrometric characterization of the FMRFamide-related peptide family in the pericardial organ of Cancer borealis

Ma¹,

Sturm²,

Kutz-Naber³

et al. 2009

Biochemical and Biophysical Research Communications

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The tetrapeptide, FMRFamide, was first discovered in 1977 in the molluscan nervous system and was found to affect the contractile force of molluscan cardiac muscle and other muscles [1]. Since then, numerous FMRFamide-related peptides (FaRPs) have been reported in both invertebrate and vertebrate species [2][3][4][5][6][7][8][9]. We have previously reported the detection and identification of numerous FaRPs in Cancer borealis pericardial organs (POs), one of the major neurosecretory structures in the crustaceans [2][3]. Here, we have developed two immunoaffinity-based methods, immunoprecipitation (IP) and immuno-dot blot screening assay, for the enrichment of FaRPs in C. borealis POs. A combined mass spectrometry (MS)-based approach involving both matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS) and nanoscale liquid chromatography coupled to electrospray ionization quadrupole time-of-flight tandem mass spectrometry (nanoLC-ESI-QTOF MS/MS) is used for a more comprehensive characterization of the FaRP family by utilizing high mass accuracy measurement and efficient peptide sequencing. Overall, 17 FMRFamide-related peptides were identified using these two complementary immunobased approaches. Among them, three novel peptides were reported for the first time in this study.

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A strategy for isolating rare peptides: isolation and sequencing of a large peptide present in a single neuron of the nematode Ascaris suum

Cited by 7 publications

References 15 publications

Molecular characterization and cell-specific expression of an ion transport peptide in the tobacco hornworm, Manduca sexta

Molecular characterization and cell-specific expression of an ion transport peptide in the tobacco hornworm, Manduca sexta

Untitled

Immunoaffinity-based mass spectrometric characterization of the FMRFamide-related peptide family in the pericardial organ of Cancer borealis

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