Search PubMed⌕ Search

Biomedical subjects

J A Hoffmann

Publications and source records attributed to J A Hoffmann.

At least 91 records · Page 5Linked to original sources
PubMed1992

Two-dimensional 1H NMR study of recombinant insect defensin A in water: resonance assignments, secondary structure and global folding.

A 500 MHz 2D 1H NMR study of recombinant insect defensin A is reported. This defense protein of 40 residues contains 3 disulfide bridges, is positively charged and exhibits antibacterial properties. 2D NMR maps of recombinant defensin A were fully assigned and secondary structure elements were localized. The set of NOE connectivities, 3JNH-alpha H coupling constants as well as 1H/2H exchange rates and delta delta/delta T temperature coefficients of NH protons strongly support the existence of an alpha-helix (residues 14-24) and of an antiparallel beta-sheet (residues 27-40). Models of the backbone folding were generated by using the DISMAN program and energy refined by using the AMBER program. This was done on the basis of: (i) 133 selected NOEs, (ii) 21 dihedral restraints from 3JNH-alpha H coupling constants, (iii) 12 hydrogen bonds mostly deduced from 1H/2H exchange rates or temperature coefficients, in addition to 9 initial disulfide bridge covalent constraints. The two secondary structure elements and the two bends connecting them involve approximately 70% of the total number of residues, which impose some stability in the C-terminal part of the molecule. The remaining N-terminal fragment forms a less well defined loop. This spatial organization, in which a beta-sheet is linked to an alpha-helix by two disulfide bridges and to a large loop by a third disulfide bridge, is rather similar to that found in scorpion charybdotoxin and seems to be partly present in several invertebrate toxins.

Amino Acid Sequence↗
PubMed1992

Insect defensins: inducible antibacterial peptides.

In response to bacterial challenge or trauma, insects produce a battery of bactericidal or bacteriostatic molecules with a broad spectrum of activity against Gram-positive and/or Gram-negative bacteria; most are small-sized cationic peptides. This review focuses on insect defensins, a large group of inducible antibacterial peptides that are present both in ancient and recent insect orders. This immune response of insects shares many of the characteristics of the mammalian acute phase response.

Amino Acid Sequence↗
PubMed1992

The effect of pancreatic polypeptide on glucose disposal after surgical alterations of the pancreas.

Surgical alterations of the pancreas result in anatomic changes that can affect postoperative glucose metabolism. Pancreas transplantation results in reduction of beta-cell mass, systemic release of insulin, and denervation. The authors hypothesized that such alterations affect peripheral glucose disposal to induce an "insensitivity" to endogenously (systemically) released insulin. Additionally, they hypothesized that surgically induced deficiency of the postprandial hormone, pancreatic polypeptide, might contribute to altered glucose disposal. The authors studied two surgical models in dogs known to be devoid of pancreatic polypeptide--70% proximal pancreatectomy (PPx) and PPx plus distal pancreas autotransplantation (PAT/B). Oral glucose challenge and euglycemic hyperinsulinemic clamp studies were performed before and after a 16-day "pulsed" infusion of pancreatic polypeptide. Both surgical procedures resulted in elevations in the integrated glucose response after oral glucose, which was not affected by pancreatic polypeptide infusion. Euglycemic clamp studies showed decreased hepatic glucose output (Ra) and overall glucose disposal (Rd) in the fasted state for both surgical groups. The transplant animals demonstrated significant decreases in Rd during the hyperinsulinemic challenge (3.2 +/- 0.01 versus 5.7 +/- 0.01 mg/kg/minute at 60 to 120 minutes for PAT/B versus control). After 16 days of pancreatic polypeptide infusion, however, basal Ra, as well as basal and 60- to 120-minute Rd values, were returned to control values in the transplant group. The authors conclude that pancreas transplantation results in altered glucose disposal, possibly due to an altered effectiveness of systemically released insulin. They conclude that pancreatic polypeptide is an important modulator of peripheral insulin action. Therefore, the role of pancreatic polypeptide must be taken into account when evaluating postoperative glucose metabolism in canine models of pancreas transplantation.

Animals↗
PubMed1991

Insect immunity. Isolation from a coleopteran insect of a novel inducible antibacterial peptide and of new members of the insect defensin family.

Injection of heat-killed bacteria into larvae of the large tenebrionid beetle Zophobas atratus (Insecta, Endopterygota, Coleoptera) results in the appearance in the hemolymph of a potent antibacterial activity as evidenced by a plate growth inhibition assay. We have isolated three peptides (A-C) from this immune hemolymph which probably account for most of this activity. Their primary structures were established by a combination of peptide sequencing and molecular mass determination by mass spectrometry. Peptide A, which is bactericidal against Gram-negative cells, is a 74-residue glycine-rich molecule with no sequence homology to known peptides. We propose the name coleoptericin for this novel inducible antibacterial peptide. Peptides B and C are isoforms of a 43-residue peptide which contains 6 cysteines and shows significant sequence homology to insect defensins, initially reported from dipteran insects. This peptide is active against Gram-positive bacteria. The results are discussed in connection with recent studies on inducible antibacterial peptides present in the three other major orders of the endopterygote clade of insects: the Lepidoptera, Diptera, and Hymenoptera.

Amino Acid Sequence↗
PubMed1991

Isolation and structural characterization of an insulin-related molecule, a predominant neuropeptide from Locusta migratoria.

Neurohaemal lobes of corpora cardiaca of Locusta migratoria are an established storage site for neurohormones produced by the neurosecretory cells of the brain. As previously reported [Hietter, H., Van Dorsselaer, A., Green, B., Denoroy, L., Hoffmann, J.A. & Luu, B. (1990) Eur. J. Biochem. 187, 241-247], the isolation and characterization of a novel 5-kDa peptide from these lobes served as the basis for oligonucleotide screening of cDNA libraries prepared from poly(A) RNA from neurosecretory cells of the central nervous system. From subsequent cDNA cloning studies [Lagueux, M., Lwoff, L., Meister, M., Goltzené, F. & Hoffmann, J.A. (1990) Eur. J. Biochem. 187, 249-254], the existence of a 145-residue precursor protein was deduced, which contained, in addition to the 5-kDa peptide, amino-acid sequences with homology to the A and B chains of an insulin-related peptide. In the present study we have isolated the native molecule from corpora cardiaca of Locusta and characterized, by Edman degradation and plasma-desorption mass spectrometry, the two chains as follows: A chain, Gly-Val-Phe-Asp-Glu-Cys-Cys-Arg-Lys-Ser-Cys-Ser-Ile-Ser-Glu-Leu-Gln-Thr- Tyr-Cys - Gly (Ile, isoleucine); B chain, Ser-Gly-Ala-Pro-Gln-Pro-Val-Ala-Arg-Tyr-Cys-Gly-Glu-Lys-Leu-Ser-Asn-Ala- Leu-Lys - Leu-Val-Cys-Arg-Gly-Asn-Tyr-Asn-Thr-Met-Phe. Taken in conjunction with the previous cloning studies, our data lead to a clear picture of the processing of Locusta preproinsulin. They indicate that locusta corpora cardiaca contain remarkably large amounts of one single insulin form, in contrast to multiple insulin isoforms of Bombyx mori, the only other insect species from which insulin-related peptides have been isolated and characterized [Nagasawa, H., Kataoka, H., Isogai, A., Tamura, S., Suzuki, A., Mizoguchi, A., Fujiwara, Y., Suzuki, A., Takahashi, S. & Ishizaki, H. (1986) Proc. Natl Acad. Sci. USA 83, 5840-5843].

Amino Acid Sequence↗
PubMed1991

Determination of disulfide bridges in natural and recombinant insect defensin A.

The primary-structure comparison of natural insect defensin A from Phormia terranovae and recombinant insect defensin A from Saccharomyces cerevisiae has been accomplished using a combination of Edman degradation and liquid secondary ion mass spectrometry. The natural and recombinant proteins have the same primary structure with identical disulfide-bond designations (formula; see text) as determined from the peptides obtained after thermolysin digestion. The combined use of Edman degradation and mass spectometry allowed the disulfide-bridge structure to be determined with a total of only 40 micrograms (9.9 nmol) natural peptide. Mass spectrometry provides a rapid means of disulfide-bridge verification, requiring not more than 20 micrograms recombinant insect defensin A, which is compatible with use in batch analysis.

Amino Acid Sequence↗
PubMed1991

Structural requirements of pancreatic polypeptide receptor binding.

Pancreatic polypeptide (PP) receptors have been identified and characterized on the basolateral membranes (BLM) of canine intestinal mucosa. The present study was designed to ascertain the structural requirements of the PP molecule for binding to its receptor. A radioreceptor assay using purified BLM was employed to elucidate receptors specific to PPs of various mammalian species and to modified bovine PP (bPP) fragments. Receptor cross-reactivities (CR) to various PPs and bPP fragments were established. Results show that percent receptor CR by PPs of various species was as follows: bPP (100%) greater than human PP (68%) greater than porcine PP (50%) greater than canine PP (45%) greater than ovine PP (36%) greater than rat PP (3%). The fragments bPP-(1-15), bPP-(1-17), bPP-(1-26), bPP-(16-23), bPP-(18-30), bPP-(24-36), bPP-(27-35), and bPP-(31-36) at 500 nM did not significantly displace tracer from receptor (less than 0.1% CR). Des-COOH-terminal tyrosinamide [bPP-(1-35)] produced less than 0.1% CR. Oxidation of bPP methionine-30 residue to methionine sulfoxide decreased displacement to 67%. Modification of native amidated tyrosinamide to the free acid abolished receptor binding, whereas esterification to the methyl ester of COOH-terminal tyrosine restored binding to 60%. Additionally, percent CR decreased progressively as amino acid residues were deleted from the NH2-terminal region. We conclude that the molecular homologue of PP primary structure is necessary for full receptor binding. Both the NH2- and COOH-terminal residues are required for recognition, and the COOH-terminal tyrosinamide must be intact for PP binding to its receptor.

Amino Acid Sequence↗
PubMed1990

Insect immunity. Characterization of a Drosophila cDNA encoding a novel member of the diptericin family of immune peptides.

Drosophila shows an immune response when challenged by injection of low doses of bacteria. To date, the molecules involved in this immune reaction have remained elusive, with the exception of cecropins (4-kDa antibacterial peptides initially isolated from the moth Hyalophora cecropia) for which three closely related genes have been characterized recently. We report the molecular cloning and sequencing of a cDNA from a library of immune Drosophila which encodes a novel member of the family of diptericins (9-kDa antibacterial peptides initially isolated from the fly Phormia terranovae). Transcripts for the Drosophila diptericin are detected 2 h after injection of bacteria. They are apparently derived from a single gene mapping at position 56 A on the right arm of the second chromosome. We discuss the existence of a distant relationship between the diptericins and two other groups of anti-bacterial insect proteins, the attacins, and the sarcotoxins II.

Amino Acid Sequence↗
PubMed1990

cDNAs from neurosecretory cells of brains of Locusta migratoria (Insecta, Orthoptera) encoding a novel member of the superfamily of insulins.

From neurohaemal lobes of corpora cardiaca of Locusta migratoria a 5-kDa peptide has been isolated and its sequence established [see the accompanying paper, by Hietter et al. (1990) Eur. J. Biochem. 187, 241-247]. We have designed oligonucleotide probes from the peptide sequence of this molecule and screened a library prepared from mRNA of the neurosecretory cell region of the brain of this insect. Several positive cDNAs were isolated, the combined nucleotide sequences of which predict a large precursor of 145 residues (15770 Da) containing the newly isolated 5-kDa peptide. The peptide is flanked by regions homologous to the A and B chains of the superfamily of insulins. The overall organization of the precursor is as follows: signal peptide/domain homologous to the B chain of insulins/C (connecting)-peptide (corresponding to the newly isolated 5-kDa peptide)/domain homologous to the A chain of insulins. The numbers and relative positions of the cysteines of the Locusta peptide are equivalent to those of the other members of the insulin superfamily and most of the hydrophobic core residues are conserved.

Amino Acid Sequence↗
PubMed1990

Insect immunity: expression of the two major inducible antibacterial peptides, defensin and diptericin, in Phormia terranovae.

Injections of low doses of bacteria into larvae of Phormia terranovae induce the appearance of potent bactericidal peptides in the blood, among which predominate the anti-Gram positive insect defensins and the anti-Gram negative diptericins. Insect defensins show significant homologies to mammalian (including human) microbicidal peptides present in polymorphonuclear leukocytes and macrophages. We report the molecular cloning of cDNAs and primer extension studies which indicate that insect defensin is produced as a prepro-peptide yielding mature defensin A (40 residues) after cleavage of a putative signal peptide (23 residues) and a prosequence (34 residues). Previous studies have established that diptericin (82 residues) is matured from a pre-peptide by cleavage of a putative signal peptide (19 residues) and C-terminal amidation. Using oligonucleotide probes complementary to the sequences of the mRNAs for defensin and diptericin, we show by in situ hybridization that both antibacterial peptides are concomitantly synthesized by the same cells: thrombocytoids, a specialized blood cell type, and adipocytes. Transcriptional studies based on hybridization of RNAs to cDNAs of defensin and diptericin indicate that the transcription of both genes is induced regardless of the nature of the stimulus (injection of Gram positive or Gram negative bacteria, lipopolysaccharides). Even a sterile injury applied to axenically raised larvae is efficient in inducing the transcription of both genes suggesting that the local disruption of the integument aspecifically initiates a signalling mechanism which the thrombocytoids and the adipocytes are able to interpret. The transcription of immune genes is relatively short lived and a second challenge yields a response similar to that of the first stimulus, indicating that the experimental insects do not keep a 'memory' of their first injection.

Amino Acid Sequence↗
PubMed1990

Purification and analysis of the major components of chum salmon protamine contained in insulin formulations using high-performance liquid chromatography.

A simple high-performance liquid chromatographic method has been developed for the rapid purification and analysis of protamine components contained in insulin formulations. Only a single step is needed to separate peptides whose compositions, sizes, and unusual isoelectric points (pI 13.8) are nearly identical. The method involves their isocratic separation on a reversed-phase column using a pH 2 phosphate buffer and a low acetonitrile content as an eluant. The purified chum salmon components were analyzed by amino acid analysis, solid-phase amino acid sequencing, carboxypeptidase B digests, insulin complexation analysis, and a mass spectrophotometric procedure which gives an accurate mass of the intact peptides. This HPLC purification technique may also be applicable to protamines and other highly basic peptides isolated from other sources.

Amino Acid Sequence↗
PubMed1989

L-canavanine incorporation into vitellogenin and macromolecular conformation.

L-Canavanine is a potentially deleterious arginine antimetabolite whose toxicity is expressed in canavanine-sensitive organisms ranging from viruses to humans. Canavanine, a substrate for arginyl-tRNA synthetase, is incorporated into nascent polypeptide chains in place of arginine. This substitution results in the production of structurally aberrant, canavanyl proteins. Chemical, physical, and immunological studies of native and canavanine-containing vitellogenin obtained from female migratory locusts (Locusta migratoria migratorioides (Orthoptera] provide the first experimental evidence that canavanine can disrupt the tertiary and/or quaternary structure that yields the three-dimensional conformation unique to the protein. These findings enhance our understanding of the biochemical basis for canavanine's antimetabolic and potent insecticidal properties.

Animals↗
PubMed1989

Insect immunity. Isolation of cDNA clones corresponding to diptericin, an inducible antibacterial peptide from Phormia terranovae (Diptera). Transcriptional profiles during immunization.

We have previously isolated and characterized a family of novel 8-kDa cationic antibacterial peptides synthesized by larvae of Phormia terranovae (Diptera) in response to various injuries. These molecules have been named diptericins. The peptide sequence of diptericin A was used to prepare oligonucleotides for screening cDNA libraries and we report in the present paper the isolation of several cDNA clones encoding diptericin. The analysis of the nucleotide sequences indicates that diptericin is synthesized as a prepeptide which is matured in two steps: (a) cleavage of a signal peptide and (b) amidation of the C-terminal residue. Interestingly, the 3' untranslated region of the mRNA contains a consensus sequence TTATTTAT which is also observed in the mRNA of another insect antibacterial peptide (attacin-related sarcotoxin IIA) and in mRNAs encoding proteins related to the inflammatory response in mammals. Our data illustrate that diptericins form a polymorphic family of immune peptides. The transcription of the diptericin genes is rapidly induced in the fat body after inoculation of bacteria, as evidenced by the transcriptional profile.

Amino Acid Sequence↗
PubMed1988

Insect immunity. Purification and characterization of a family of novel inducible antibacterial proteins from immunized larvae of the dipteran Phormia terranovae and complete amino-acid sequence of the predominant member, diptericin A.

Injury or injection of live bacteria into third instar larvae of the dipteran insect Phormia terranovae results in the appearance in the haemolymph of at least five groups of heat-stable, more or less basic peptides with antibacterial activity against Escherichia coli. Three of these peptides have been purified. The amino acid sequence has been completely established for one of these and partially (first 40 residues from the N-terminus) for the two others. The sequences show marked homologies indicating that the three peptides belong to a common family. They are not related to other known antibacterial peptides from insects [lysozymes, cecropins (including sarcotoxin I) and attacins]. We propose the name of diptericins for this new family of antibiotic molecules.

Amino Acid Sequence↗
PubMed1988

Further experimental evidence for the involvement of ecdysone in the control of meiotic reinitiation in oocytes of Locusta migratoria (Insecta, Orthoptera).

Ecdysone has recently been shown to be able to trigger meiotic reinitiation in vitro in submature oocytes of Locusta. In the present study we have experimentally depressed (by 60-70%) ecdysone biosynthesis in the ovaries of adult females by rearing them on a diet with a modified sterol profile. Mature oocytes from such females fail to undergo normal reinitiation, but when incubated in vitro, can be induced to break their meiotic arrest by the addition of exogenous ecdysone. These results lend further support to the hypothesis that in Locusta, ovarian ecdysone is involved in the control of meiotic reinitiation.

Animals↗
PubMed1988

Characterization of three hydroxylases involved in the final steps of biosynthesis of the steroid hormone ecdysone in Locusta migratoria (Insecta, Orthoptera).

It is most generally accepted that the last three enzymatic reactions in the biosynthetic pathway of ecdysone are, in this order, the hydroxylations at positions C-25, C-22 and C-2. Using high specific activity tritiated ecdysone precursors (2,22,25-trideoxyecdysone, 2,22-dideoxyecdysone and 2-deoxyecdysone) we have characterized the hydroxylases involved in these reactions, in the major biosynthetic tissue of ecdysone, i.e. the prothoracic glands. We show that C-2 hydroxylase is a mitochondrial oxygenase which differs from conventional cytochrome P-450-dependent monooxygenases by its relative insensitivity to CO. In contrast, C-22 and C-25 hydroxylases appear as classical cytochrome P-450 monooxygenases; C-22 hydroxylase is a mitochondrial enzyme whereas our data point to a microsomal localization of the C-25 hydroxylase.

Animals↗
PubMed1987

Ecdysteroid biosynthesis and embryonic development are disturbed in insects (Locusta migratoria) reared on plant diet (Triticum sativum) with a selectively modified sterol profile.

Wheat seedlings germinating in the presence of the systemic fungicide fenpropimorph accumulate 9beta,19-cyclopropylsterols (95% of total sterols) in place of Delta(5)-sterols, which are normally produced in these plants. Adult females of the phytophagous insect Locusta migratoria show a dramatic decrease in their cholesterol content when reared on fenpropimorph-treated wheat. These females lay eggs with the ecdysteroid concentration reduced by up to 80% as compared to controls. Injection of fenpropimorph to the insects or feeding them on wheat coated with the fungicide (normal sterol composition) does not affect their sterol or ecdysteroid profiles; addition of cholesterol to fenpropimorph-treated wheat prior to feeding restores normal ecdysteroid titers in the insects. The severe reduction of the ecdysteroid content in eggs laid by females reared on fenpropimorph-treated wheat is associated with a series of developmental arrests and/or abnormalities. The results show that the dietary 9beta,19-cyclopropylsterols cannot be used by Locusta in place of Delta(5)-sterols for ecdysteroid biosynthesis. They suggest that the selective inhibition of specific enzymes in the sterol biosynthetic pathway of the plants can be used as a strategy to control insect development.

Journal Article↗