scholarly journals ISOLATION AND IDENTIFICATION BY SEQUENCE ANALYSIS OF EXPERIMENTALLY INDUCED GUINEA PIG AMYLOID FIBRILS

1974 ◽  
Vol 140 (3) ◽  
pp. 871-876 ◽  
Author(s):  
Martha Skinner ◽  
Edgar S. Cathcart ◽  
Alan S. Cohen ◽  
Merrill D. Benson
Keyword(s):  
Amino Acid ◽  
Sequence Analysis ◽  
Guinea Pig ◽  
Guinea Pigs ◽  
Amyloid Fibrils ◽  
Protein A ◽  
Terminal Sequence ◽  
Isolation And Identification ◽  
Amyloid Deposits ◽  
Immunologic Analysis

Amyloidosis was produced experimentally in guinea pigs by multiple casein injections. Amyloid fibrils were isolated and fractionated and a protein obtained that had an amino acid composition comparable with A protein, a unique nonimmunoglobulin constituent of secondary amyloid deposits. N-terminal sequence analysis demonstrated a sequence homologous with that of A proteins from human and monkey preparations but preceded by a 5-residue peptide which had an N-terminal histidine. A definite species specificity in A protein from human and guinea pig was identified on immunologic analysis.

scholarly journals FURTHER STUDIES ON THE ROLE OF PROTEASES IN THE ALLERGIC REACTION

1961 ◽  
Vol 113 (2) ◽  
pp. 359-380 ◽  
Author(s):  
Georges Ungar ◽  
Takuso Yamura ◽  
Jacqueline B. Isola ◽  
Sidney Kobrin
Keyword(s):  
Amino Acid ◽  
Guinea Pig ◽  
Proteolytic Activity ◽  
Guinea Pigs ◽  
Protease Activity ◽  
Amino Acid Esters ◽  
Protein Substrates ◽  
Protease Activation ◽  
Acid Esters

Protease activity was measured through the hydrolysis of synthetic amino acid esters in body fluids and tissues of guinea pigs, rats, mice, and humans. Significant in vitro activation was observed in serum and lung slices of sensitized guinea pigs on addition of the specific antigen. Increased proteolytic activity was also seen in reverse anaphylaxis. More marked activation occurred when guinea pig serum was treated with peptone and guinea pig or rat serum was treated with agar. Protease activation was demonstrated in specimens of human skin under the influence of a poison ivy extract or croton oil added in vitro. Urinary protease activity of guinea pigs increased significantly during the first hours of anaphylactic shock and very markedly in peptone shock. Peptone shock, elicited in mice pretreated with H. pertussis, was accompanied by a considerable increase in protease activity in the peritoneal fluid as compared with non-pretreated mice which were insensitive to peptone. Proteolytic activity resulting from the activation procedures was due to a number of proteases. The dominant substrate affinity and inhibition patterns suggest that serum and urine proteases are similar to but not identical with plasmin. Anaphylactic activation exhibited patterns different from those resulting from the action of anaphylactoid agents. Tissue enzymes are either of cathepsin- or chymotrypsin-type or mixtures of both. Some of the activated enzymes, although remarkably effective in hydrolyzing amino acid esters, show no activity on protein substrates. This does not justify, however, their designation as "esterases." They probably belong to the class of specific proteases acting only on a single or a small number of functionally significant protein substrates. There is at present sufficient evidence to prove not only that protease activation does occur in anaphylaxis and anaphylactoid conditions but also that it is an important component of the chain of reactions leading to the allergic response.

SOME PROPERTIES OF APOFERRITIN ISOLATED FROM GUINEA PIG LIVER

1962 ◽  
Vol 40 (1) ◽  
pp. 983-987 ◽  
Author(s):  
Felix Friedberg
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Guinea Pig ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Guinea Pigs ◽  
Dicarboxylic Acids ◽  
Acetate Buffer ◽  
Pig Liver ◽  
Guinea Pig Liver

Apoferritin isolated from livers of guinea pigs and characterized by a s°w,20 of 17.7 and a pI of 4.8 (in acetate buffer Γ/2 0.1) was hydrolyzed with 5.7 N HCl for 22 and 44 hours and its amino acid composition determined. The protein appears rich in dicarboxylic acids and in leucine. The content of sulphur-containing amino acids is fairly small.

scholarly journals Clinical and Ultrastructural Studies of Gelatinous Drop-Like Corneal Dystrophy (GDLD) of a Patient with TACSTD2 Gene Mutation

2019 ◽  
Vol 2019 ◽  
pp. 1-7
Author(s):  
Ali Masmali ◽  
Aljoharah Alkanaan ◽  
Hind M. Alkatan ◽  
Omar Kirat ◽  
Abdullah Ayidh Almutairi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amyloid Fibrils ◽  
Polarized Light ◽  
Corneal Dystrophy ◽  
Corneal Tissue ◽  
Ultrastructural Studies ◽  
Amyloid Deposits ◽  
Exon 1 ◽  
Pas Staining ◽  
The Right

Purpose. To describe clinical, molecular genetics, histopathologic and ultrastructural findings of gelatinous drop-like corneal dystrophy (GDLD) (OMIM #204870) in a Sudanese patient. Method. An ocular examination revealed the onset of GDLD in a Sudanese patient (50 years old) at King Khalid Specialist Hospital, Riyadh. The 333 sequence variants in 13 GDLD genes of a DNA sample were screened by Asper Ophthalmics Ltd. It was further confirmed by sequencing. The patient had undergone a penetrating keratoplasty in the right eye. The corneal tissue was processed for histopathology and ultrastructural studies. Results. Slit-lamp observation showed grayish-white multiple superficial corneal nodules of various sizes in the left and right eye. Both corneas became clear after the surgery. The GDLD deposits in the subepithelial region and in the anterior stroma were confirmed by PAS staining and their apple-green birefringence under polarized light. Ultrastructurally, the amyloid fibrils were very thin and grouped in whorl-like structures, which caused splits between and within the stromal lamellae. Collagen fibrils (CFs) and keratocytes had degenerated. A homozygous c.355T > A mutation in exon 1 of the TACSTD2 (M1S1) gene was detected, and alteration of the amino acid (p.Cysl19Ser in NCBI entry NP_002344.2) was observed. Conclusion. In our patient with GDLD, a “c.355T > A” mutation in exon 1 of TACSTD2 was detected and believed to be responsible for the alteration of the amino acid leading to the formation of the amyloid deposits. The deposits caused the ultrastructural degeneration of epithelium, Bowman’s layer, stroma, and keratocytes of the GDLD cornea.

scholarly journals Processing of the precursor of protamine P2 in mouse. Peptide mapping and N-terminal sequence analysis of intermediates

1991 ◽  
Vol 277 (1) ◽  
pp. 39-45 ◽  
Author(s):  
D Carré-Eusèbe ◽  
F Lederer ◽  
K H D Lê ◽  
S M Elsevier
Keyword(s):  
Amino Acid ◽  
Sequence Analysis ◽  
Peptide Mapping ◽  
Chromatin Condensation ◽  
Peptide Bond ◽  
Chromosomal Protein ◽  
Amino Acid Residues ◽  
Terminal Sequence ◽  
Vinylidene Difluoride ◽  
N Terminus

Protamine P2, the major basic chromosomal protein of mouse spermatozoa, is synthesized as a precursor almost twice as long as the mature protein, its extra length arising from an N-terminal extension of 44 amino acid residues. This precursor is integrated into chromatin of spermatids, and the extension is processed during chromatin condensation in the haploid cells. We have studied processing in the mouse and have identified two intermediates generated by proteolytic cleavage of the precursor. H.p.l.c. separated protamine P2 from four other spermatid proteins, including the precursor and three proteins known to possess physiological characteristics expected of processing intermediates. Peptide mapping indicated that all of these proteins were structurally similar. Two major proteins were further purified by PAGE, transferred to poly(vinylidene difluoride) membranes and submitted to automated N-terminal sequence analysis. Both sequences were found within the deduced sequence of the precursor extension. The N-terminus of the larger intermediate, PP2C, was Gly-12, whereas the N-terminus of the smaller, PP2D, was His-21. Both processing sites involved a peptide bond in which the carbonyl function was contributed by an acidic amino acid.

THROMBOSPONDIN INTERACTION WITH PLASMINOGEN

1987 ◽  
Author(s):  
Patricia DePoli ◽  
Theresa Bacon-Baquley ◽  
Daniel A Walz
Keyword(s):  
Amino Acid ◽  
Sequence Analysis ◽  
Terminal Sequence ◽  
Independent Manner ◽  
Amino Terminal ◽  
High Affinity ◽  
Distinct Site ◽  
Tissue Plasminogen ◽  
Trimolecular Complex ◽  
Amino Terminal Sequence

Platelet thrombospondin (TSP) interacts with plasminogen in a specific and saturable manner. TSP can form a trimolecular complex with histidine-rich glycoprotein and plasminogen and the plasminogen within such complexes can reportedly be activated by tissue plasminogen activator. We have studied the interaction of TSP with plasminogen using Western blotting of plasminogen, reduced plasmin and the elastase-generated fragments of plasminogen and their binding of iodinated TSP. TSP was found to specifically bind to plasminogen and the heavy (non-enzyme) chain of plasmin in a calcium-independent manner. Binding could be blocked by preincubation of the immobilized plasminogen or plasmin with an excess of unlabeled TSP. Plasminogen domains (kringles) were generated by limited eTastase proteolysis. TSP bound specifically to a single 51 kDa plasminogen fragment. The elastase-generated fragments were separated by lysine-Sepharose chromatography and their identities established by amino acid composition and amino-terminal sequence analysis. The 51 kDa plasminogen fragment bound to lysine-Sepharose and had an amino-terminal sequence corresponding to kringle 4 (K4) and a composition consistent with that of K4-K5-plasmin. TSP binding to this fragment was not blocked by the presence of an excess of the fragment K1-K2-K3, K4, nor miniplasminogen (K5-plasmin). Binding does not appear to be directly dependent upon the specific high-affinity lysine binding site of the 51 kDa fragment. Our data suggests that thrombospondin interacts with plasminogen at a single distinct site, and that this recognition site is at or near the K4-K5 contiguous region of plasminogen.

scholarly journals Protein A Suppresses Immune Responses during Staphylococcus aureus Bloodstream Infection in Guinea Pigs

mBio ◽  
2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Hwan Keun Kim ◽  
Fabiana Falugi ◽  
Lena Thomer ◽  
Dominique M. Missiakas ◽  
Olaf Schneewind
Keyword(s):  
Staphylococcus Aureus ◽  
Guinea Pig ◽  
B Cell ◽  
Bloodstream Infection ◽  
Guinea Pigs ◽  
Protective Immunity ◽  
Protein A ◽  
Pig Model ◽  
Content Type ◽  
Guinea Pig Model

ABSTRACT  Staphylococcus aureusinfection is not associated with the development of protective immunity, and disease relapses occur frequently. We hypothesize that protein A, a factor that binds immunoglobulin Fcγ and cross-links VH3 clan B cell receptors (IgM), is the staphylococcal determinant for host immune suppression. To test this, vertebrate IgM was examined for protein A cross-linking. High VH3 binding activity occurred with human and guinea immunoglobulin, whereas mouse and rabbit immunoglobulins displayed little and no binding, respectively. Establishing a guinea pig model of S. aureus bloodstream infection, we show that protein A functions as a virulence determinant and suppresses host B cell responses. Immunization with SpAKKAA, which cannot bind immunoglobulin, elicits neutralizing antibodies that enable guinea pigs to develop protective immunity.IMPORTANCE Staphylococcus aureusis the leading cause of soft tissue and bloodstream infections; however, a vaccine with clinical efficacy is not available. Using mice to model staphylococcal infection, earlier work identified protective antigens; however, corresponding human clinical trials did not reach their endpoints. We show that B cell receptor (IgM) cross-linking by protein A is an important immune evasion strategy of S. aureus that can be monitored in a guinea pig model of bloodstream infection. Further, immunization with nontoxigenic protein A enables infected guinea pigs to elicit antibody responses that are protective against S. aureus. Thus, the guinea pig model may support preclinical development of staphylococcal vaccines.

scholarly journals Molecular cloning of cDNA encoding an unrecognized component of amyloid in Alzheimer disease

1993 ◽  
Vol 90 (23) ◽  
pp. 11282-11286 ◽  
Author(s):  
K Uéda ◽  
H Fukushima ◽  
E Masliah ◽  
Y Xia ◽  
A Iwai ◽  
...  
Keyword(s):  
Amino Acid ◽  
Alzheimer Disease ◽  
Amyloid Fibrils ◽  
Protein A ◽  
Microscopic Analysis ◽  
Amino Acid Sequences ◽  
Full Length ◽  
Peptide Sequence ◽  
Electron Microscopic ◽  
Hydrophobic Domain

A neuropathological hallmark of Alzheimer disease (AD) is a widespread amyloid deposition. We analyzed the entire amino acid sequences in an amyloid preparation and found, in addition to the major beta/A4-protein (A beta) fragment, two unknown peptides. We raised antibodies against synthetic peptides using subsequences of these peptides. These antibodies immunostained amyloid in neuritic and diffuse plaques as well as vascular amyloid. Electron microscopic analysis demonstrated that the immunostaining was localized on amyloid fibrils. We have isolated an apparently full-length cDNA encoding a 140-amino-acid protein within which two previously unreported amyloid sequences are encoded in tandem in the most hydrophobic domain. We tentatively named this 35-amino acid peptide NAC (non-A beta component of AD amyloid) and its precursor NACP. NAC is the second component, after A beta, identified chemically in the purified AD amyloid preparation. Secondary structure predictions indicate that the NAC peptide sequence has a strong tendency to form beta-structures consistent with its association with amyloid. NACP is detected as a M(r) 19,000 protein in the cytosolic fraction of brain homogenates and comigrates on immunoblots with NACP synthesized in Escherichia coli from NACP cDNA. NACP mRNA is expressed principally in brain but is also expressed in low concentrations in all tissues examined except in liver, suggesting its ubiquitous and brain-specific functions. The availability of the cDNA encoding full-length NACP should help to elucidate the mechanisms of amyloidosis in AD.

scholarly journals Calcitonin and chromogranin A localization in medullary carcinoma of the thyroid by immunoelectron microscopy.

1988 ◽  
Vol 36 (8) ◽  
pp. 1031-1036 ◽  
Author(s):  
M M Silver ◽  
S A Hearn ◽  
L D Lines ◽  
M Troster
Keyword(s):  
Tumor Cells ◽  
Immunoelectron Microscopy ◽  
Chromogranin A ◽  
Amyloid Fibrils ◽  
Secretory Granules ◽  
Protein A ◽  
Medullary Carcinoma ◽  
Amyloid Deposits ◽  
Microscopy Method ◽  
Rat Thyroid

We used a post-embedding immunoelectron microscopy method, using protein A-gold, to detect calcitonin and chromogranin A immunoreactivity in three cases of human medullary thyroid carcinoma. Because the epoxy-embedded tissue had been fixed (glutaraldehyde or formaldehyde) and osmicated before embedment, the proteins were identified in optimally preserved tissue. Uranyl and lead staining was used after immunolabeling, so that the tissue was also optimally contrasted. The morphological advantage provided by osmication was tested by labeling rat thyroid gland C-cells for calcitonin. The protein A-gold technique allowed localization of both antigens to the contents of membrane-bound secretory granules in the tumor cells. In one case, labeling density for each antigen was measured over several intercellular compartments and the interstitium. Calcitonin, but not chromogranin A, reactivity was also identified in intracellular amyloid fibrils in two cases, showing that the constant region of calcitonin is preserved in amyloid deposits related to the tumor cells.

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