scholarly journals A STUDY OF IMMUNOGLOBULIN STRUCTURE

1966 ◽  
Vol 124 (3) ◽  
pp. 307-330 ◽  
Author(s):  
C. Baglioni ◽  
D. Cioli
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Peptide Mapping ◽  
Gel Filtration ◽  
Bence Jones Protein ◽  
Type K ◽  
The Common ◽  
L Proteins ◽  
Peptide Maps ◽  
Bence Jones Proteins

Urinary proteins of patients with myeloma, prepared by precipitation with ammonium sulphate, have been separated by gel filtration on Sephadex G-100 after reduction and aminoethylation. Many specimens separated into a major peak of Bence Jones protein and into minor peaks of albumin, a protein tentatively identified with heavy chain and a smaller molecular weight protein corresponding to the variable portion of the corresponding Bence Jones protein. The Bence Jones protein purified by gel filtration was analyzed by electrophoresis and by peptide mapping after tryptic digestion. The peptide maps of 24 type K and 20 type L Bence Jones proteins were compared. A set of common peptides was identified in the peptide maps of the Bence Jones proteins of the same type; the common peptides of type K proteins were completely different from the common peptides of type L proteins. The patterns of distinctive peptides was compared; no similarities were found between distinctive peptides of type K and of type L proteins. Some similarities were observed in the distinctive peptides of proteins of the same type. The similarities involved in many cases peptides containing cysteine, whereas similarities in other peptides were limited. This observation suggested that the amino acid sequence around the cysteines of the variable NH2-terminal half of the Bence Jones proteins may show less variability than other sequences. A few proteins of the same type differed in all their distinctive peptides, an indication that multiple amino acid differences exist between individual Bence Jones proteins. The genetic mechanisms responsible for the variability in the amino acid sequence of the NH2-terminal half of the light chains of immunoglobulins are discussed in view of the results of the comparison by peptide mapping of the Bence Jones proteins.

scholarly journals A STUDY OF IMMUNOGLOBULIN STRUCTURE

1969 ◽  
Vol 130 (2) ◽  
pp. 401-415 ◽  
Author(s):  
R. Quattrocchi ◽  
D. Cioli ◽  
C. Baglioni
Keyword(s):  
Light Chain ◽  
Peptide Mapping ◽  
Gel Filtration ◽  
Antigenic Determinants ◽  
Minimal Size ◽  
Bence Jones Protein ◽  
Peptide Map ◽  
Peptide Maps ◽  
Similar Peptide ◽  
Bence Jones Proteins

102 human Bence Jones proteins have been purified by gel filtration, digested with trypsin, and analyzed by peptide mapping. In several cases Bence Jones "fragments", corresponding to the variable half of the corresponding proteins, were observed. The peptide maps of the proteins were compared to establish whether any identical proteins were present in the sample analyzed. No Bence Jones protein showed a peptide map identical to that of any other protein, although remarkable similarities in the peptide maps were observed for some proteins. Two proteins that gave very similar peptide maps were then examined in detail, by purifying and analyzing the tryptic peptides. It was then found that these two proteins differ in amino acid sequence in at least six positions. The probability of not finding two identical sequences by examining a sample extracted from populations of light chains of different sizes has been calculated. This has led to an estimate of the minimal size of the population of light chain sequences in humans. The number of light chain sequences appears to be at least a few thousand. Information on the frequency of Inv and Oz antigenic determinants and on the relative frequency of subtypes of K chains has been obtained. Proteins of KI subtype are found most frequently. The possibility that different subtypes may be predominant in different species is discussed in relation to the evolutionary arguments used in favor of the somatic theories on the origin of variability of immunoglobulin chains.

Chemical structure of light chains: amino acid sequence of type K Bence-Jones proteins

1966 ◽  
Vol 166 (1003) ◽  
pp. 124-137 ◽  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Sequence Data ◽  
Single Point ◽  
Structural Difference ◽  
Light Chains ◽  
Single Point Mutation ◽  
Amino Terminal ◽  
Type K ◽  
Bence Jones Proteins

Bence-Jones proteins are the light chains of the autologous myeloma globulin and are analogous to the light chains of normal human immunoglobulins. Peptide mapping has demonstrated that Bence-Jones proteins share a fixed portion of their sequence (the ‘constant’ portion) and also have a mutable part (the ‘variable’ portion). Sequence analysis and ordering of the tryptic and chymotryptic peptides has provided the tentative complete amino acid sequence of one Bence-Jones protein of antigenic type K. Comparison with partial sequence data for other type K Bence-Jones proteins has revealed many structural differences in the amino terminal half of the molecules, but only one structural difference in the carboxyl terminal half. The latter is strongly correlated with the Inv genetic factor. The points of interchange in the amino terminal half occur in clusters close to the half cystine residues and the ‘switch peptide’ (positions 102 through 105), after which the sequence becomes essentially invariant. This suggests that the major areas subject to sequence variation are part of a single topographical region which may define a portion of the antigen combining site in the light chains of antibodies. Many, but not all, the amino acid interchanges are compatible with a single point mutation. As yet, no single mutational theory suffices to explain the manifold differences in structure of the light chains. Such structural variation, however, could result from the presence of many related genes.

scholarly journals CATABOLIC ORIGIN OF A BENCE JONES PROTEIN FRAGMENT

1968 ◽  
Vol 128 (3) ◽  
pp. 517-532 ◽  
Author(s):  
D. Cioli ◽  
C. Baglioni
Keyword(s):  
Amino Acids ◽  
Free Amino Acids ◽  
Free Amino ◽  
Peptide Mapping ◽  
Gel Filtration ◽  
Urinary Proteins ◽  
Small Peptides ◽  
Protein Fragment ◽  
Bence Jones Protein ◽  
Bence Jones Proteins

Gel filtration analysis of the urinary proteins of some patients with myeloma has shown the presence of "fragments" of Bence Jones proteins which correspond to the variable half of these proteins. Experiments have been carried out to establish the origin of a "fragment" observed in a patient who excreted a large amount of this protein. Labeled homologous Bence Jones protein has been injected into this and other control patients. Excretion of labeled "fragment" has been observed in all. Analysis by peptide mapping and radio-autography of this labeled "fragment" isolated from the urine showed that the invariable half of the Bence Jones protein was not excreted; it seemed thus likely that the invariable half was metabolized to small peptides and free amino acids. A labeled Bence Jones protein which was excreted without any accompanying "fragment" was injected into the patient who excreted large amounts of "fragment." No excretion of labeled "fragment" was observed. It was thus concluded that the property of being degraded to "fragment" is characteristic of some "fragile" Bence Jones proteins and is not determined by the patient. Incubation with serum or urine of the "fragile" Bence Jones protein failed to produce any "fragment." "Fragments" of Bence Jones proteins are thus most likely formed during excretion of these proteins through the kidney and are products of the catabolism of Bence Jones proteins.

scholarly journals Purification, characterization and cDNA cloning of an endo-exonuclease from the basidiomycete fungus Armillaria mellea

1999 ◽  
Vol 339 (3) ◽  
pp. 713-720
Author(s):  
Vincent HEALY ◽  
Shawn DOONAN ◽  
Tommie V. McCARTHY
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Cdna Cloning ◽  
Peptide Mapping ◽  
Gel Filtration ◽  
Significant Degree ◽  
Prolonged Incubation ◽  
Armillaria Mellea ◽  
Cloned Cdna ◽  
Basidiomycete Fungus

We have purified an endo-exonuclease from the fruiting body of the basidiomycete fungus Armillaria mellea by using an ethanol fractionation step, followed by two rounds of column chromatography. The enzyme had an apparent molecular mass of 17500 Da and was shown to exist as a monomer by gel-filtration analysis. The nuclease was active on both double-stranded and single-stranded DNA but not on RNA. It was optimally active at pH 8.5 and also exhibited a significant degree of thermostability. Three bivalent metal ions, Mg2+, Co2+ and Mn2+, acted as cofactors in the catalysis. It was also inhibited by high salt concentrations: activity was completely abolished at 150 mM NaCl. The nuclease possessed both endonuclease activity on supercoiled DNA and a 3ʹ-5ʹ (but not a 5ʹ-3ʹ) exonuclease activity. It generated 5ʹ-phosphomonoesters on its products that, after a prolonged incubation, were hydrolysed to a mixture of free mononucleotides and small oligonucleotides ranging in size from two to eight bases. Elucidation of its N-terminal amino acid sequence permitted the cDNA cloning of the A. mellea nuclease via a PCR-based approach. Peptide mapping of the purified enzyme generated patterns consistent with the amino acid sequence coded for by the cloned cDNA. A BLAST search of the SwissProt database revealed that A. mellea nuclease shared significant amino acid similarity with two nucleases from Bacillussubtilis, suggesting that the three might constitute a distinct class of nucleolytic enzymes.

scholarly journals Immunoglobulin λ-chains. The complete amino acid sequence of a Bence-Jones protein

1968 ◽  
Vol 110 (4) ◽  
pp. 631-652 ◽  
Author(s):  
C. Milstein ◽  
J. B. Clegg ◽  
J. M. Jarvis
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Total Amino Acid ◽  
Cysteine Residues ◽  
Bence Jones Protein ◽  
Chromatographic Techniques ◽  
Peptic Digestion ◽  
Similarities And Differences ◽  
Methionine Residues ◽  
Bence Jones Proteins

The total amino acid sequence of a λ Bence-Jones protein has been established. The protein contains 211 residues, which include two methionine residues. Splitting with cyanogen bromide gave three fragments, the largest of which included the C-terminal half, which is common to other Bence-Jones proteins of the same type. The peptides obtained by tryptic, chymotryptic and peptic digestion were isolated and purified by paper-electrophoretic and chromatographic techniques. Reduction followed by carboxymethylation of the cysteine residues with radioactive iodoacetate was found to be a powerful tool in the isolation of some insoluble peptides. Unusual features of the molecule are the fact that it contains six cysteine residues and not five as observed in both κ and λ Bence-Jones proteins studied previously, and its size, which seems two residues smaller than the smallest Bence-Jones protein studied hitherto. The similarities and differences between this and other Bence-Jones proteins are discussed.

scholarly journals The Amino Acid Sequence of a κ Type Bence-Jones Protein

1969 ◽  
Vol 244 (13) ◽  
pp. 3537-3549 ◽  
Author(s):  
E J Whitley ◽  
K Titani ◽  
F W Putnam
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Bence Jones Protein

scholarly journals The Amino Acid Sequence of a κ Type Bence-Jones Protein

1969 ◽  
Vol 244 (13) ◽  
pp. 3521-3536 ◽  
Author(s):  
K Titani ◽  
E J Whitley ◽  
F W Putnam
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Bence Jones Protein

scholarly journals The Amino Acid Sequence of a κ Type Bence-Jones Protein

1969 ◽  
Vol 244 (13) ◽  
pp. 3550-3560 ◽  
Author(s):  
K Titani ◽  
T Shinoda ◽  
F W Putnam
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Bence Jones Protein

scholarly journals Streptococcus salivarius Fimbriae Are Composed of a Glycoprotein Containing a Repeated Motif Assembled into a Filamentous Nondissociable Structure

2001 ◽  
Vol 183 (9) ◽  
pp. 2724-2732 ◽  
Author(s):  
Céline Lévesque ◽  
Christian Vadeboncoeur ◽  
Fatiha Chandad ◽  
Michel Frenette
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Cell Surface ◽  
Polyclonal Antibodies ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Spontaneous Mutant ◽  
Streptococcus Salivarius ◽  
Streptococcal Species ◽  
Repeated Motif

ABSTRACT Streptococcus salivarius, a gram-positive bacterium found in the human oral cavity, expresses flexible peritrichous fimbriae. In this paper, we report purification and partial characterization of S. salivarius fimbriae. Fimbriae were extracted by shearing the cell surface of hyperfimbriated mutant A37 (a spontaneous mutant of S. salivarius ATCC 25975) with glass beads. Preliminary experiments showed that S. salivariusfimbriae did not dissociate when they were incubated at 100°C in the presence of sodium dodecyl sulfate. This characteristic was used to separate them from other cell surface components by successive gel filtration chromatography procedures. Fimbriae with molecular masses ranging from 20 × 106 to 40 × 106Da were purified. Examination of purified fimbriae by electron microscopy revealed the presence of filamentous structures up to 1 μm long and 3 to 4 nm in diameter. Biochemical studies of purified fimbriae and an amino acid sequence analysis of a fimbrial internal peptide revealed that S. salivarius fimbriae were composed of a glycoprotein assembled into a filamentous structure resistant to dissociation. The internal amino acid sequence was composed of a repeated motif of two amino acids alternating with two modified residues: A/X/T-E-Q-M/φ, where X represents a modified amino acid residue and φ represents a blank cycle. Immunolocalization experiments also revealed that the fimbriae were associated with a wheat germ agglutinin-reactive carbohydrate. Immunolabeling experiments with antifimbria polyclonal antibodies showed that antigenically related fimbria-like structures were expressed in two other human oral streptococcal species, Streptococcus mitis andStreptococcus constellatus.

Physicochemical properties of a novel α-L-arabinofuranosidase fromRhizomucor pusillusHHT-1

2001 ◽  
Vol 47 (8) ◽  
pp. 767-772 ◽  
Author(s):  
A KM Shofiqur Rahman ◽  
Shinya Kawamura ◽  
Masahiro Hatsu ◽  
M M Hoq ◽  
Kazuhiro Takamizawa
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Mass ◽  
Gel Filtration ◽  
Hydrolytic Activity ◽  
Exchange Chromatography ◽  
Ph Optimum ◽  
Rhizomucor Pusillus ◽  
Terminal Amino ◽  
Metal Cofactors

The zygomycete fungus Rhizomucor pusillus HHT-1, cultured on L(+)arabinose as a sole carbon source, produced extracellular α-L-arabinofuranosidase. The enzyme was purified by (NH4)2SO4fractionation, gel filtration, and ion exchange chromatography. The molecular mass of this monomeric enzyme was 88 kDa. The native enzyme had a pI of 4.2 and displayed a pH optimum and stability of 4.0 and 7.0–10.0, respectively. The temperature optimum was 65°C, and it was stable up to 70°C. The Kmand Vmaxfor p-nitrophenyl α-L-arabinofuranoside were 0.59 mM and 387 µmol·min–1·mg–1protein, respectively. Activity was not stimulated by metal cofactors. The N-terminal amino acid sequence did not show any similarity to other arabinofuranosidases. Higher hydrolytic activity was recorded with p-nitrophenyl α-L-arabinofuranoside, arabinotriose, and sugar beet arabinan; lower hydrolytic activity was recorded with oat–spelt xylan and arabinogalactan, indicating specificity for the low molecular mass L(+)-arabinose containing oligosaccharides with furanoside configuration.Key words: α-L-arabinofuranosidase, enzyme purification, amino acid sequence, Rhizomucor pusillus.

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