Purification and properties of manducin, an amino acid storage protein of the haemolymph of larval and pupal Manduca sexta

Abbreviation: CBP, Cerura biliproteinRunning title: Hexameric arylphorin-type insect biliprotein 2 SummaryThe abundant blue hemolymph protein of last instar larvae of the moth Cerura vinula was purified and characterized by protein-analytical, spectroscopic and electron microscopic methods. Amino acid sequences obtained from a large number of cleavage peptides revealed high similarity of the blue protein with arylphorins from a number of other moth species. In particular there is a high abundance of the aromatic amino acids tyrosine and phenylalanine amounting to about 19% of total amino acids, and a low content of methionine (0.8%) in the Cerura protein. The mass of the native protein complex was studied

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“…18,35 The CBP complex did not dissociate under these conditions, like the arylphorin (manducin) of tobacco hornworm, Manduca sexta. 36 …”

Section: Discussionmentioning

confidence: 99%

Isolation, Characterisation and Molecular Imaging of a High-Molecular-Weight Insect Biliprotein, a Member of the Hexameric Arylphorin Protein Family

Kayser

Mann

Machaidze

et al. 2009

Journal of Molecular Biology

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“…L5d7) a total of 2.5 ml haem olym ph was collected from the surviving four larvae. The proce dure for the isolation of m anducin was with m inor modifications the same as described previously [5] (see also [13]): chromatography on Sepharose 6-B and combining the protein containing fractions of Mt range 200 000-500 000; freezing at -20 °C for 24 h; thawing and centrifugation (15 m in; 3600 x^; 4 °C). The supernatant was 50% saturated with ammonium sulfate and stirred magnetically for 6 h at 4°C .…”

Section: Preparation O F [ ( U-hc)-l-tyrosine]manducinmentioning

confidence: 99%

Incorporation of Radiolabeled Tyrosine, N-Acetyldopamine, N-β-Alanyldopamine, and the Arylphorin Manducin into the Sclerotized Cuticle of Tobacco Hornworm (Manduca sexta) Pupae

Grün

Peter

1984

Zeitschrift Für Naturforschung C

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Sclerotization of the cuticle depends in many insects on the oxidation of 1 ,2 -diphenolic substrates in the outer parts of the integument. Reactive intermediates such as o-quinones, /?-quinonemethides or free radicals, which may be generated during the oxidative transformations are thought to effect covalent crosslinking of structural proteins by Michael-type additions and/or free radical poly merizations (for a recent review see [1]). Chem ical models are apparently consistent with such theories. However, structures of crosslinks in sclerotized cuticle are essentially unknown due to the difficulties encountered in attempts to isolate soluble deriv atives of native structures from the exoskeleton. Thus, an unambiguous chemical identification of a crosslink partial structure has not been reported with full details. Likewise, very little is known on the identity of structural polypeptides participating in the formation of crosslinks.Applications of immunological [2], radiochem ical [2,3], and histochemical (K. Scheller, personal

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“…Fractions that induced significant increases in numbers of cultured stem cells were subsequently analyzed by Edman degradation (Blackburn et al, 2004). An active peptide of Mr 77 KDa was identified; its sequence aligned nicely with the known sequence of Manduca sexta a-arylphorin (Kramer et al, 1980;Blackburn et al, 2004). The arylphorins are high molecular weight (approximately 450,000 KDa) complexes composed of six subunits of a and b forms (Kramer et al, 1980).…”

Section: Ecdysonesmentioning

confidence: 59%

“…An active peptide of Mr 77 KDa was identified; its sequence aligned nicely with the known sequence of Manduca sexta a-arylphorin (Kramer et al, 1980;Blackburn et al, 2004). The arylphorins are high molecular weight (approximately 450,000 KDa) complexes composed of six subunits of a and b forms (Kramer et al, 1980). Although they are expressed primarily by the fat body of late instar larvae, they are released into the larval hemolymph, where they become the major protein component, and later accumulate in the fat body (Webb and Riddiford, 1988).…”

Section: Ecdysonesmentioning

confidence: 60%

“…3), stimulated H. virescens midgut stem cells to multiply and continually form clusters of new cells after the 3rd to 4th day of continuous exposure, with as little as 125 ng/ml in the culture medium (Blackburn et al, 2004). Previously, it was postulated that the large titers of arylphorin synthesized by caterpillars about to enter metamorphosis served as storage protein, a source of amino acids for new tissues (Kramer et al, 1980;Webb and Riddiford, 1988). However, we showed that a-arylphorin could act as a powerful growth factor that stimulated stem cell mitosis, providing cells for new or enlarged tissues in subsequent stages of development.…”

Section: Ecdysonesmentioning

confidence: 99%

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Factors affecting proliferation and differentiation of lepidopteran midgut stem cells

Loeb

2010

Arch Insect Biochem Physiol

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Midgut stem cells of last instar larvae and pupae of Heliothis virescens, Lymantria dispar and several other Lepidopteran species have been cultured in vitro and have been induced to proliferate using low titers of ecdysteroids and the 77-Kda peptide fragment, a-arylphorin, isolated and identified from pupal fat body tissue. The insulin-related hormone, Bombyxin, also induced mitosis in cultured midgut stem cells; it appeared to be fast-acting and quickly inactivated, while a-arylphorin was slower to act and had a longer lasting effect in vitro, indicating different functions for these proliferation agents. Changes in Calcium ion concentration within or outside the cells discretely affected stem cell differentiation, indicating a role for second messenger participation in peptide regulation of this process. Four different peptides (MDFs 1-4) that induced midgut stem cells to differentiate to mature midgut cell types in vitro were isolated and characterized from conditioned media and hemolymph of H. virescens and L. dispar. However, platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and all-trans retinoic acid (RA) from vertebrate sources induced differentiation to nonmidgut cell types as well. MDF1 was located in basal areas of columnar cells of midgut epithelium, although MDF2 was observed in all of the cytoplasm of columnar cells and in droplets of antibody positive material in the midgut lumen, suggesting a digestive function as well for this Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA and does not imply its approval to the exclusion of other products that may also be suitable. Correspondence to: Marcia J. Loeb, 6920 Fairfax Rd.,

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Purification and properties of manducin, an amino acid storage protein of the haemolymph of larval and pupal Manduca sexta

Cited by 120 publications

References 21 publications

Isolation, Characterisation and Molecular Imaging of a High-Molecular-Weight Insect Biliprotein, a Member of the Hexameric Arylphorin Protein Family

Isolation, Characterisation and Molecular Imaging of a High-Molecular-Weight Insect Biliprotein, a Member of the Hexameric Arylphorin Protein Family

Incorporation of Radiolabeled Tyrosine, N-Acetyldopamine, N-β-Alanyldopamine, and the Arylphorin Manducin into the Sclerotized Cuticle of Tobacco Hornworm (Manduca sexta) Pupae

Factors affecting proliferation and differentiation of lepidopteran midgut stem cells

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