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PubMed2005

Helix-turn-helix peptides that form alpha-helical fibrils: turn sequences drive fibril structure.

Models of apolipoprotein A-I (apo A-I), the main protein of high-density lipoprotein, predict that it contains 10 amphiphilic, alpha-helical segments connected by turns. We synthesized four peptides with two identical 18-residue, amphiphilic, alpha-helical segments (Anantharamaiah, G. M., et al. (1985) J. Biol. Chem. 260, 10248-10255) connected by putative turn sequences from apo A-I: (1) Ac-DWLKAFYDKVAEKLKEAFKVEPLRADWLKAFYDKVAEKLKEAF-NH2, (2) Ac-DWLKAFYDKVAEKLKEAFGLLPVLEDWLKAFYDKVAEKLKEAF-NH2, (3) Ac-DWLKAFYDKVAEKLKEAFKVQPYLDDWLKAFYDKVAEKLKEAF-NH2, and (4) Ac-DWLKAFYDKVAEKLKEAFNGGARLADWLKAFYDKVAEKLKEAF-NH2. Surprisingly, peptides 1-3 formed fibrils after incubation (37 degrees C, 10 mM sodium phosphate, pH 7.60), but in contrast to beta-sheet amyloid fibrils, these did not bind thioflavin T and they induced a blue shift in the spectrum of Congo red. CD (peptides 1-3) and FTIR (peptides 1 and 2) of the fibrils showed significant alpha-helical character. Synchrotron X-ray fiber diffraction on a magnetically aligned sample of 1 confirmed the alpha-helical character in the fibrils and indicated that the helical axes are oriented perpendicular to the fibril axis. In contrast, peptide 4, containing two Gly residues but no Pro in the turn, formed only a small amount of nonfibrillar precipitate after prolonged incubation. Peptide 4P (peptide 4 with a Pro in place of the central Ala) and peptide 5, containing a PEG block in lieu of the central turn, were similar to peptide 4 in not forming fibrils, possibly because the region linking the helices was unstructured. These studies indicate that varying turn sequences between longer amphiphilic alpha-helical segments can drive the structure of fibrils.

Amino Acid Sequence↗
PubMed2000

Peripheral nerve amyloidosis in sural nerve biopsies: a clinicopathologic analysis of 13 cases.

OBJECTIVE: Amyloidosis is a well-recognized but uncommon cause of peripheral neuropathy. Our objectives were to determine the overall prevalence of peripheral nerve amyloidosis in sural nerve biopsies and to evaluate the clinical and pathologic features of these lesions. METHODS: All available histologic and ultrastructural materials on biopsy tissue from 13 cases of peripheral nerve amyloidosis were examined. Muscle biopsies performed at the same time as the nerve biopsy were reviewed when available. Clinical data were collected on all patients. RESULTS: The prevalence of amyloidosis in sural nerve biopsies at our institution was 13 (1.2%) of 1098 cases over a 15.8-year period. These patients ranged in age from 41 to 82 years (median, 61 years) at initial presentation and included 10 men and 3 women. Presenting neuropathy symptoms were sensory in 6 of the 13 patients, motor in 2 cases, and mixed in 5 cases. Cardiac, renal, or gastrointestinal involvement was present in 7 of 13 cases. Two patients had myeloma and 7 had systemic autonomic symptoms. Two patients had probable familial amyloid polyneuropathy, and 1 patient demonstrated an alanine 60 point mutation. Amyloid, identified as amorphous eosinophilic extracellular deposits demonstrating apple green birefringence on Congo red stain or recognized by its characteristic fibrillar ultrastructure by electron microscopy, was identified in the endoneurium in 12 nerves, perineurium in 2 nerves, and epineurium in 9 nerves. Chronic inflammation was identified in 5 nerves. Axonal loss was recorded as mild (<25%) in 1 nerve, moderate (25% to 75%) in 8 nerves, and severe (>75%) in 4 nerves. Axonal degeneration predominated over demyelination in 8 of 10 cases that could be evaluated. Concomitant muscle biopsies contained amyloid deposits in 8 of 9 cases. CONCLUSIONS: Amyloidosis is a rare (1.2% in our series) cause of peripheral neuropathy with a distinct microscopic and ultrastructural appearance. Just over half the patients in our study had visceral organ involvement and systemic autonomic symptoms. The peripheral neuropathy was associated with axonal degeneration and a moderate to severe axonal loss in the majority of cases. Amyloid deposition was present in 8 out of 9 muscle biopsies performed at the same time.

Adult↗
PubMed2003

A systematic investigation into the effect of protein destabilisation on beta 2-microglobulin amyloid formation.

Beta-2-microglobulin (beta(2)m) has been shown to form amyloid fibrils with distinct morphologies under acidic conditions in vitro. Short, curved fibrils (<600 nm in length), form rapidly without a lag phase, with a maximum rate at pH 3.5. By contrast, fibrils with a long (approximately 1 microm), straight morphology are produced by incubation of the protein at pH< or =3.0. Both fibril types display Congo red birefringence, bind Thioflavin-T and have X-ray fibre diffraction patterns consistent with a cross-beta structure. In order to investigate the role of different partially folded states in generating fibrils of each type, and to probe the effect of protein stability on amyloid formation, we have undertaken a detailed mutagenesis study of beta(2)m. Thirteen variants containing point mutations in different regions of the native protein were created and their structure, stability and fibril forming propensities were investigated as a function of pH. By altering the stability of the native protein in this manner, we show that whilst destabilisation of the native state is important in the generation of amyloid fibrils, population of specific denatured states is a pre-requisite for amyloid formation from this protein. Moreover, we demonstrate that the formation of fibrils with different morphologies in vitro correlates with the relative population of different precursor states.

Amyloid↗
PubMed1993

Hemin uptake in Porphyromonas gingivalis: Omp26 is a hemin-binding surface protein.

A 26-kDa outer membrane protein (Omp26) has been proposed to play a role in hemin acquisition by Porphyromonas gingivalis (T. E. Bramanti and S. C. Holt, J. Bacteriol. 174:5827-5839, 1992). We studied [55Fe]hemin uptake in P. gingivalis grown under conditions of hemin starvation (Omp26 expressed on the outer membrane surface) and hemin excess (Omp26 not expressed on surface). [55Fe]hemin uptake occurred rapidly in hemin-starved cells which incorporated up to 70% of total [55Fe]hemin within 3 min. P. gingivalis grown under hemin-starved conditions or treated with the iron chelator 2,2'-bipyridyl to induce an iron stress took up six times more [55Fe]hemin than hemin-excess-grown cells. Polyclonal monospecific anti-Omp26 antibody added to hemin-starved cells inhibited [55Fe]hemin uptake by more than 50%, whereas preimmune serum had no effect. [55Fe]hemin uptake in hemin-starved P. gingivalis was inhibited (36 to 67%) in the presence of equimolar amounts of unlabeled hemin, protoporphyrin IX, zinz protoporphyrin, and Congo red dye but was not inhibited in the presence of non-hemin-containing iron sources. Heat shock treatment (45 degrees C) of hemin-excess-grown P. gingivalis (which cases translocation of Omp26 to the surface) increased [55Fe]hemin uptake by threefold after 3 min in comparison with cells grown at 37 degrees C. However, no [55Fe] hemin uptake beyond 3 min was observed in either hemin-excess-grown or hemin-starved cells exposed to heat shock. In experiments using heterobifunctional cross-linker analysis, hemin and selected porphyrins were cross-linked to Omp26 in hemin-starved P. gingivalis, but no cross-linking was seen with hemin-excess-grown cells. However, cross-linking of hemin to Omp26 was observed after heat shock treatment of hemin-excess-grown cells. Finally, anti-Omp26 antibody inhibited cross-linked of hemin to Omp26. These findings indicate that hemin binding and transport into P.gingivalis cell mediated by Omp26.

Bacterial Outer Membrane Proteins↗
PubMed1995

Fibrillogenesis in Alzheimer's disease of amyloid beta peptides and apolipoprotein E.

A central event in Alzheimer's disease is the conformational change from normally circulating soluble amyloid beta peptides (A beta) and tau proteins into amyloid fibrils, in the form of senile plaques and neurofibrillary tangles respectively. The apolipoprotein E (apoE) gene locus has recently been associated with late-onset Alzheimer's disease. It is not know whether apoE plays a direct role in the pathogenesis of the disease. In the present work we have investigated whether apoE can affect the known spontaneous in vitro formation of amyloid-like fibrils by synthetic A beta analogues using a thioflavine-T assay for fibril formation, electron microscopy and Congo Red staining. Our results show that, under the conditions used, apoE directly promotes amyloid fibril formation, increasing both the rate of fibrillogenesis and the total amount of amyloid formed. ApoE accelerated fibril formation of both wild-type A beta-(1-40) and A beta-(1-40A), an analogue created by the replacement of valine with alanine at residue 18, which alone produces few amyloid-like fibrils. However, apoE produced only a minimal effect on A beta-(1-40Q), found in the Dutch variant of Alzheimer's disease. When recombinant apoE isoforms were used, apoE4 was more efficient than apoE3 at enhancing amyloid formation. These in vitro observations support the hypothesis that apoE acts as a pathological chaperone, promoting the beta-pleated-sheet conformation of soluble A beta into amyloid fibres, and provide a possible explanation for the association of the apoE4 genetic isoform with Alzheimer's disease.

Alzheimer Disease↗
PubMed2000

Distribution of beta-amyloid and amyloid precursor protein in the brain of spawning (senescent) salmon: a natural, brain-aging model.

Brain amyloid precursor protein (APP), a normal constituent of neurons, glial cells and cerebrospinal fluid, has several proposed functions (e.g., in neuronal growth and survival). It appears, however, that altered processing of APP is an initial or downstream step in the neuropathology of brain aging, Alzheimer's disease (AD), and Down's syndrome (DS). Some studies suggest that proteolytic cleavage of APP, producing beta-amyloid (Abeta(1-42)), could have neurotoxic or neuroprotective effects. In this study, we utilized antibodies to human APP(695) and Abeta(1-42,) and Congo red staining, to search for amyloid deposition in the brain of semelparous spawning kokanee salmon (Oncorhynchus nerka kennerlyi). Intracellular APP(695) immunoreactivity (APP-ir) was observed in brain regions involved in gustation (glomerulosus complex), olfaction (putative hippocampus, olfactory bulb), vision (optic tectum), the stress response (nucleus preopticus and nucleus lateralis tuberis), reproductive behavior (nucleus preopticus magnocellularis, nucleus preopticus periventricularis, ventral telencephalon), and coordination (cerebellum). Intra- and extra-neuronal Abeta(1-42) immunoreactivity (Abeta-ir) were present in all APP-ir regions except the nucleus lateralis tuberis and Purkinje cells of the cerebellum (coordination). Thus, the relationship between APP and Abeta deposition during brain aging could shed light on the processing of APP into Abeta, neurodegeneration, and possible protection of neurons that are functioning in spawning but senescent salmon. Pacific salmon, with their predictable and synchronized life history, could provide research options not available with the existing models for studies of brain aging and amyloidosis.

Aging↗
PubMed2000

Modification of the aggregation behaviour of the environmental Ralstonia eutropha-like strain AE815 is reflected by both surface hydrophobicity and amplified fragment length polymorphism (AFLP) patterns.

After inoculation of the plasmid-free non-aggregative Ralstonia eutropha-like strain AE815 in activated sludge, followed by reisolation on a selective medium, a mutant strain A3 was obtained, which was characterized by an autoaggregative behaviour. Strain A3 had also acquired an IncP1 plasmid, pLME1, co-aggregated with yeast cells when co-cultured, and stained better with Congo red than did the AE815 strain. Contact angle measurements showed that the mutant strain was considerably more hydrophobic than the parent strain AE815, and scanning electron microscopy (SEM) revealed the production of an extracellular substance. A similar hydrophobic mutant (AE176R) could be isolated from the AE815-isogenic R. eutropha-like strain AE176. With the DNA fingerprinting technique repetitive extragenic palindromic-polymerase chain reaction (REP-PCR), no differences between these four strains, AE815, A3, AE176 and AE176R, could be revealed. However, using the amplified fragment length polymorphism (AFLP) DNA fingerprinting technique with three different primer combinations, small but clear reproducible differences between the banding patterns of the autoaggregative mutants and their non-autoaggregative parent strains were observed for each primer set. These studies demonstrate that, upon introduction of a strain in an activated sludge microbial community, minor genetic changes readily occur, which can nevertheless have major consequences for the phenotype of the strain and its aggregation behaviour.

Adaptation, Physiological↗
PubMed2004

Amyloid neuropathy: a retrospective study of 35 peripheral nerve biopsies.

We performed a retrospective study of 35 peripheral nerve biopsies (PNBs) with amyloid deposits in the endoneurium. In every case, nerve lesions were studied on paraffin-embedded fragments (PEFs) and by ultrastructural examination (USE). In addition, muscle fragments were taken and embedded in paraffin. Immunohistochemistry was performed with anti-transthyretin (TTR) serum on 19 nerve and 15 muscle PEFs. Direct immunofluorescence with anti-light-chain sera was performed on frozen nerve fragments in 19 cases. Endoneurial amyloid deposits were easily identified on routine PEF in 26 cases, after Congo red or thioflavine staining in three, and by USE in six. A dramatic myelinated fiber loss was evidenced in 34 cases (77-2970 per mm2), and features of axonal degeneration were present in every case. Segmental demyelination was observed in 10 cases. A mutation in the TTR gene was present in 14 cases, with Met30 mutation in 10 and Ala49 in four members of the same family. Amyloid deposits were strongly marked by the anti-TTR serum in 11 other cases, twice in the endoneurium, five around muscle fibers, and four in both locations. In eight patients, light-chain positivity was evidenced in endoneurial deposits, lambda in six and kappa in two. Two other patients with monoclonal gammopathy did not present any light-chain fixation. In 17 cases, amyloidosis was disclosed by PNB and 13 had a TTR pathology; eight of them, over 65 years old, correspond to a late-onset form of familial amyloid polyneuropathy which is an underdiagnosed condition.

Adult↗
PubMed2006

Microstructural imaging of early gel layer formation in HPMC matrices.

A real-time confocal fluorescence imaging method has been developed which allows the critical early stages of gel layer formation in hydroxypropylmethylcellulose (HPMC) matrices to be examined. Congo Red, a fluorophore whose fluorescence is selectively intensified when bound to beta-D-glucopyranosyl sequences, has allowed mapping of hydrated polymer regions within the emerging gel layer, and revealed for the first time, the microstructural sequence of polymer hydration during development of the early gel layer. Liquid penetration and swelling can be examined in unprecedented detail. The earliest images revealed an initial phase of liquid ingress into the tablet pore network, followed by the progressive formation of a coherent gel layer by outward columnar swelling and coalescence of hydrated HPMC particles. Salts can markedly affect HPMC matrix behaviour. Gel layer growth in 0.1-0.5 M NaCl was progressively suppressed until at 0.75 M, particles clearly failed to coalesce into a gel layer, although with considerable polymer swelling. The failure to form a limiting diffusion barrier resulted in enhanced liquid penetration of the core, and the swelling of particles that did not coalesce culminated in surface disintegration. This provides direct evidence of physical mechanisms that contribute to salts accelerating drug release from HPMC matrices.

Congo Red↗
PubMed1993

Immunolocalization of scrapie amyloid in non-congophilic, non-birefringent deposits in golden Syrian hamsters with experimental transmissible mink encephalopathy.

Transmissible mink encephalopathy (TME), a naturally occurring subacute spongiform encephalopathy in commercially ranch-reared mink (Mustela vision), is characterized neuropathologically by spongiform changes in the neuropil, intracytoplasmic neuronal vacuolation and astrocytic hypertrophy and hyperplasia. Amyloid deposits have not been observed in brain tissue sections from animals with natural and experimental TME using conventional histochemical stains such as Congo red. To determine if amyloid deposits be visualized by immunocytochemical techniques, we stained formalin-fixed, formic acid-treated brain tissue sections from several animal species with natural and experimental TME, using a rabbit antiserum directed against scrapie amyloid (PrP27-30). Scrapie amyloid-immunoreactive deposits were found in golden Syrian hamsters experimentally infected with TME, but were absent in mink with natural and experimental TME, as well as in ferrets and squirrel monkeys with experimental TME. The scrapie amyloid-immunoreactive deposits, which were non-congophilic and non-birefringent, were distributed in the subependymal, subpial and perivascular regions of the brain, as in hamsters infected with the 263K strain of scrapie. Ultrastructurally, scrapie amyloid-immunoreactive deposits revealed a collection of degenerating neurites with numerous abnormal mitochondria and degenerating synapses. Amyloid fibrils were not observed. Anti-scrapie amyloid antibodies immunoabsorbed with scrapie amyloid abolished immunostaining. Our data indicate the presence of scrapie amyloid lacking the molecular conformation of amyloid fibrils in hamsters with experimental TME.

Animals↗
PubMed1998

Sulfate content and specific glycosaminoglycan backbone of perlecan are critical for perlecan's enhancement of islet amyloid polypeptide (amylin) fibril formation.

Islet amyloidosis is characterized by the deposition and accumulation of amylin in pancreatic beta-cells and is observed in 90% of patients with type 2 diabetes. Previous studies have also revealed the presence of the specific heparan sulfate proteoglycan, perlecan, colocalized to islet amyloid deposits, similar to perlecan's known involvement with other amyloid proteins. In the present study, perlecan purified from the Engelbreth-Holm-Swarm (EHS) tumor was used to define perlecan's interactions with amylin (i.e., islet amyloid polypeptide) and its effects on amylin fibril formation. Using a solid phase-binding immunoassay, human amylin, but not rat amylin, bound immobilized EHS perlecan with a single dissociation constant (Kd) = 2.75 x 10(-6) mol/l. The binding of human amylin to perlecan was similarly observed using perlecan heparan sulfate glycosaminoglycans (GAGs), and was completely abolished by 10 micromol/l heparin. Using thioflavin T fluorometry, Congo red staining, and electron microscopy methodology, intact perlecan was found to enhance amylin fibril formation in a dosage-dependent manner, with the majority of these effects attributed to the heparan sulfate GAG chains of perlecan. Other sulfated GAGs and related macromolecules were also effective in the enhancement of amylin fibril formation in the order of heparin > heparan sulfate > chondroitin-4-sulfate = dermatan sulfate = dextran sulfate > pentosan polysulfate, implicating the importance of the specific GAG/carbohydrate backbone. The sulfate content of heparin/heparan sulfate was also important for the enhancement of amylin fibril formation in the order of heparin > N-desulfated N-acetylated heparin > completely desulfated N-sulfated heparin > completely desulfated N-acetylated heparin. These studies suggest that the enhancement effects of perlecan on amylin fibril formation are mediated primarily by both specific GAG chain backbone and GAG sulfate content, and implicate perlecan as an important macromolecule that is likely involved in the pathogenesis of islet amyloidosis.

Amyloid↗
PubMed2000

Evidence for seeding of beta -amyloid by intracerebral infusion of Alzheimer brain extracts in beta -amyloid precursor protein-transgenic mice.

Many neurodegenerative diseases are associated with the abnormal sequestration of disease-specific proteins in the brain, but the events that initiate this process remain unclear. To determine whether the deposition of the beta-amyloid peptide (Abeta), a key pathological feature of Alzheimer's disease (AD), can be induced in vivo, we infused dilute supernatants of autopsy-derived neocortical homogenates from Alzheimer's patients unilaterally into the hippocampus and neocortex of 3-month-old beta-amyloid precursor protein (betaAPP)-transgenic mice. Up to 4 weeks after the infusion there was no Abeta-deposition in the brain; however, after 5 months, the AD-tissue-injected hemisphere of the transgenic mice had developed profuse Abeta-immunoreactive senile plaques and vascular deposits, some of which were birefringent with Congo Red. There was limited deposition of diffuse Abeta also in the brains of betaAPP-transgenic mice infused with tissue from an age-matched, non-AD brain with mild beta-amyloidosis, but none in mice receiving extract from a young control case. Abeta deposits also were not found in either vehicle-injected or uninjected transgenic mice or in any nontransgenic mice. The results show that cerebral beta-amyloid can be seeded in vivo by a single inoculation of dilute AD brain extract, demonstrating a key pathogenic commonality between beta-amyloidosis and other neurodegenerative diseases involving abnormal protein polymerization. The paradigm can be used to clarify the conditions that initiate in vivo beta-amyloidogenesis in the brain and may yield a more authentic animal model of Alzheimer's disease and other neurodegenerative disorders.

Adult↗
PubMed2000

Biophysical studies of the development of amyloid fibrils from a peptide fragment of cold shock protein B.

The peptide CspB-1, which represents residues 1-22 of the cold shock protein CspB from Bacillus subtilis, has been shown to form amyloid fibrils when solutions containing this peptide in aqueous (50%) acetonitrile are diluted in water [M. Gross et al. (1999) Protein Science 8, 1350-1357] We established conditions in which reproducible kinetic steps associated with the formation of these fibrils can be observed. Studies combining these conditions with a range of biophysical methods reveal that a variety of distinct events occurs during the process that results in amyloid fibrils. A CD spectrum indicative of beta structure is observed within 1 min of the solvent shift, and its intensity increases on a longer timescale in at least two kinetic phases. The characteristic wavelength shift of the amyloid-binding dye Congo Red is established within 30 min of the initiation of the aggregation process and corresponds to one of the phases observed by CD and to changes in the Fourier transform-infrared spectrum indicative of beta structure. Short fibrillar structures begin to be visible under the electron microscope after these events, and longer, well-defined amyloid fibrils are established on a timescale of hours. NMR spectroscopy shows that there are no significant changes in the concentration of monomeric species in solution during the events leading to fibril formation, but that soluble aggregates too large to be visible in NMR spectra are present throughout the process. A model for amyloid formation by this peptide is presented which is consistent with these kinetic data and with published work on a variety of disease-related systems. These findings support the concept that the ability to form amyloid fibrils is a generic property of polypeptide chains, and that the mechanism of their formation is similar for different peptides and proteins.

Amino Acid Sequence↗
PubMed1998

When noncongophilic glomerular fibrils do not represent fibrillary glomerulonephritis: nonspecific mesangial fibrils in sclerosing glomeruli.

In addition to fibrillary glomerulonephritis (FGN), Congo red negative mesangial fibrils may commonly be seen in sclerosing glomerular diseases. Rarely, these nonspecific mesangial fibrils (NMF) may mimic fibrils in FGN and cause a differential diagnostic pitfall. Following an interesting case of sclerosing crescentic glomerulonephritis with abundant NMF (which is presented in some detail) we have reviewed our renal biopsy files for a period of two and a half years and found additional 16 cases where the presence of NMF warranted studies to exclude FGN and other diseases with fibrillary deposits. The immunofluorescence pattern characteristically seen in FGN was not present in any of these cases. Our data confirm that mesangial fibrillary material seen ultrastructurally in sclerosing glomeruli with negative or nonspecific immunofluorescence (IF) represents a nonspecific reaction of the mesangial matrix to chronic glomerular injury. The presence of NMF should not lead to the erroneous diagnosis of FGN. Negative or nonspecific immunofluoresence, localization to the mesangium in a usually segmental fashion, and the more bundle-like than random arrangement of fibrils are helpful diagnostic hints in differentiating NMF from fibrils in FGN.

Actin Cytoskeleton↗
PubMed1993

Bis-azo dyes interference with effector activation of antibodies.

Azo dyes (Congo Red, Evans Blue, Trypan Blue) were used to study the relation between antigen binding and effector activity in SRBC rabbit anti-SRBC IgG system. Antigen binding was tested in this study by agglutination and effector activity by complement fixation. Antibodies induced by the antigen binding appeared to become structurally susceptible for interaction with the dye. The dyes cause the enhancement of antigen binding by antibodies and inhibition of their effector activity. The intensity of both effects caused by binding the dye seems correlated. The increase of antibody concentration makes the effector activity of immune complexes more resistant to dye inhibition indicating that the influence of antigen binding may be exerted on assembling of antibody in the immune complex rather than on structural modulation of Fc. Also the analysis of dye effects on antibody derivatives deprived of effector activity (Fab)2, IgG with split interheavy chain disulphides supplied evidence that the induced by the antigen long distance structural effects may play the role of immune signal by influencing the assembling of antibodies and stability of immune complex.

Agglutination Tests↗
PubMed2003

A catalogue of molecular, physiological and symbiotic properties of soybean-nodulating rhizobial strains from different soybean cropping areas of China.

We have analysed 198 fast-growing soybean-nodulating rhizobial strains from four different regions of China for the following characteristics: generation time; number of plasmids; lipopolysaccharide (LPS), nodulation factors (LCOs) and PCR profiles; acidification of growth medium; capacity to grow at acid, neutral, and alkaline pH; growth on LC medium; growth at 28 and 37 degrees C; melanin production capacity; Congo red absorption and symbiotic characteristics. These unbiased analyses of a total subset of strains isolated from specific soybean-cropping areas (an approach which could be called "strainomics") can be used to answer various biological questions. We illustrate this by a comparison of the molecular characteristics of five strains with interesting symbiotic properties. From this comparison we conclude, for instance, that differences in the efficiency of nitrogen fixation or competitiveness for nodulation of these strains are not apparently related to differences in Nod factor structure.

Bacterial Proteins↗
PubMed1995

In vitro fibril formation from alpha 1-antitrypsin-derived C-terminal peptides.

Fragments from various proteolytically degraded precursor proteins can form beta-amyloid fibrils. We studied, by electron microscopy and quantitative Congo red binding, the ability of three synthetic peptides, corresponding to residues 359-374 (C-36), 370-374 (C-5) and 375-394 (C-20) from the C-terminal part of alpha 1-antitrypsin (AAT) to form beta-amyloid fibrils in vitro. The peptides C-36 and C-5 had a pronounced tendency to form fibrils. C-20 lacked this property, suggesting that residues 359-375 and/or 370-374 are most critical for fibril formation. Native AAT added to peptide 125I-C-36 could bind and form complexes with the peptide, resulting in inhibition of amyloid fibril formation. Moreover, native AAT added to preformed fibrils induced disaggregation of fibrillar structures. The structural rearrangements of AAT that occurred during this 'autointeraction' included polymerization of the serpin, and an increase of its thermal stability. Also, following interaction, an increase (20-40%) of AAT's antielastase activity was noted. The demonstration of an in vitro beta-amyloid fibril formation from the AAT derived C-terminal peptides C-36 and C-5 and its regulation by the intact AAT molecule may have important in vivo implications.

Amino Acid Sequence↗
PubMedDate not supplied

Fine needle aspiration of abdominal fat for the diagnosis of amyloidosis.

Amyloidosis is a dysproteinemia characterized by extracellular deposition of amyloid fibrils. Its diagnosis depends on the demonstration of the characteristic apple-green birefringence in tissue stained with Congo red. Aspiration of subcutaneous fat as a means of procuring tissue has received attention in the internal medicine literature. However, this application of fine needle aspiration biopsy has not been investigated extensively by cytopathologists. We therefore report our experience. Over the past two years we performed 18 aspirations of subcutaneous fat on 17 patients in whom amyloidosis was clinically suspected. Eight aspirates were positive for amyloid, and nine were negative. There were no known false-negative results. False positives were more problematic, occurred early in our experience and were due to overinterpretation of occasional long, slender, green strands representing collagen. The true positives were all characterized by large numbers of short, apple-green strands intimately associated with the fat, oriented in multiple directions and occurring in much, if not all, of the aspirated fat. Attention to details of the aspirated material other than the presence or absence of apple-green birefringence can lead to a correct distinction between amyloid and collagen. We conclude that abdominal fat pad aspiration is useful in the workup of suspected amyloidosis, especially since it is a safe, easily performed procedure.

Abdomen↗