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Biomedical subjects

M Fougereau

Publications and source records attributed to M Fougereau.

At least 73 records · Page 4Linked to original sources
PubMed1983

Localization of the two free thiol groups in the porcine pancreatic alpha-amylase I sequence.

Porcine pancreatic alpha-amylase I, a single 496 residue long polypeptide chain, contains 5 disulfide bridges and 2 free -SH groups. The conditions for specific blocking of native amylase either with radioactive N-ethyl maleimide or with labeled iodoacetic acid were determined. Under these conditions 2 moles of blocking reagent are incorporated per mole of amylase. [14C]-S-succinimido amylase was cleaved by CNBr and the resulting peptides were purified. Only one of them the CNBr 2 + 3 peptide (178 residues) was found labeled. Ts1 a 33-residue peptide containing the whole radioactivity was purified from the tryptic digest of this large fragment. After reduction and carboxymethylation Ts1A, (22 residues) was obtained which contains 2 moles of succinyl-Cys and one mole of CM-Cys per mole of peptide. Chymotryptic digestion of Ts1A yielded 2 equally labeled peptides: C1 (16 residues) and C2 (6 residues). Automated sequencing of both peptides and counting of the PTH-amino acids shows that the free cysteines are only 15 residues apart in the sequence.

Amino Acid Sequence↗
PubMed1983

Analysis of antibody diversity: V-D-J mRNA nucleotide sequence of four anti-GAT monoclonal antibodies. A paucigene system using alternate D-J recombinations to generate functionally similar hypervariable regions.

The nucleotide sequence of four anti-(Glu60-Ala30-Tyr10)n (GAT) monoclonal gamma 1 heavy chain mRNAs was determined from codon 10 to 120. This sequence overlaps with the NH2-terminal amino acid sequence, allowing elucidation of the complete protein sequence encompassing regions VH, D and JH. These sequences, which are highly conserved, indicate that anti-GAT antibodies expressing the same public idiotypic specificities represent a paucigene system, which uses at least two D-J combinations leading to functionally similar hypervariable regions involved in the recognition of the dominant Glu-Tyr determinant. D regions are encoded by D genes which are closely related either to the D-SP2 or the D.FL16 germ line gene cores.

Amino Acid Sequence↗
PubMed1983

Interstrain conservation of the murine GAT-specific antibody V kappa repertoire as analyzed at the germline gene level.

A cDNA library was constructed in pBR322 from mRNA encoding an anti-GAT (Glu60 Ala30 Tyr10) monoclonal antibody kappa chain. Two cDNA clones were extensively characterized. One, L XI 62, was derived from an aberrant V kappa-J kappa rearrangement which resulted in a frame-shift at position 96, leading to a stop codon at the very beginning of the constant region. The second, L XIX 27, 1150 bp long, was unequivocally assigned to a GAT-specific kappa chain, by comparison of its nucleotide sequence with the previously determined NH2-terminal amino acid sequence of the isolated kappa chain. A specific probe, containing the leader and most of the V kappa gene-encoded region, was prepared from this clone and hybridized to EcoRI and BamHI restriction fragments of liver (unrearranged) DNA extracted from the BALB/c, DBA/2 and C57BL/6 mouse strains. Under stringent conditions, similar patterns were observed for all three strains, and consisted of a small number of bands (3-5). Under nonstringent conditions, patterns were again very similar when the different strains were compared, although 15-20 bands could be identified. These observations support the hypothesis that the GAT-specific kappa chains found in antibodies expressing the public CGAT idiotypes are encoded by a very small number of germline genes. This V kappa repertoire seems extremely conserved between the three strains that were analyzed, an observation which correlates with the interstrain conservation of these public idiotypic specificities.

Alanine↗
PubMed1983

Structural correlates to the rabbit immunoglobulin heavy chain a100 allotype.

Three a100/a100 homozygous rabbits immunized with Micrococcus lysodeikticus produced large amounts of anti-polysaccharide antibodies of restricted heterogeneity. These antibodies were purified by either immunoabsorption or ion exchange chromatography. The almost complete sequence of one heavy chain spanning residues 1-123, with the exception of 10 residues (66-67 and 79-86), was determined. Partial sequence data were also obtained for the two other heavy chains. The identity of these three sequences in the first framework region unraveled a prototype sequence of the a100 allotype that differs from homologous sequence of VH regions that determine other allotypic specificities. The gradient of sequence conservation was found to be a100 greater than a3 greater than a1 greater than a- greater than a2. Homologies in sequence paralleled previously described serological cross-reactions observed between a100, a3 and a1 specificities. This remarkable conservation of framework residues suggests that the VH regions of the rabbit immunoglobulins represent a paucigene system, in which each basic allotypic specificity might be encoded a discrete subgroup of genes.

Amino Acid Sequence↗
PubMed1983

Structural bases for public idiotypic specificities of monoclonal antibodies directed against poly(Glu60Ala30Tyr10) and poly(Glu60Ala40) random copolymers.

NH2-terminal amino acid sequences of heavy and light chains of seven poly(Glu60Ala30Tyr10) (GAT) specific hybridoma products derived from DBA/2 and (DBA/2 X BALB/c)F1 hybrid mice and those of BALB/B polyclonal antibodies have been determined over the first 40 residues. Comparison of these sequences with those of nine other GAT or poly(Glu60Ala40) (GA) specific hybridoma products previously reported allowed the following conclusions. (i) Sequences of hybridoma H and L chains are present in the pool of polyclonal antibodies. (ii) The public CGAT (or pGAT) idiotypic specificities are strictly confined to antibodies exhibiting limited heterogeneity with regard to both the variable heavy (VH) and the variable kappa (V kappa) sequences that may be accounted for by one and two germ-line genes, respectively. (iii) The public idiotypic specificities GA-1, expressed by some anti-GAT and most anti-GA antibodies, make use of the same (or similar) VH germ-line genes as the CGAT or pGAT antibodies but possess a distinctive V kappa sequence. (iv) Antibodies expressing neither of the alternative public specificities mentioned above appear to be more heterogeneous and express VH and V kappa sequences that were found to differ from the basic structures defining the CGAT (pGAT) or GA-1 correlates. It is concluded that CGAT (or pGAT) and GA-1 public idiotypic specificities are germ-line markers of both VH and V kappa regions, an observation in agreement with previously reported serological data.

Animals↗
PubMed1982

The limited diversity of the mouse gamma-chains anti-GAT repertoire does not seem to be noticeably amplified upon class switch.

NH2-terminal sections of H and L chains isolated from five monoclonal anti-GAT antibodies derived from BALB/c mice have been sequenced upon to residue 43. Four among these five antibodies, sharing similar public idiotypic determinants, possess extremely conserved sequences, both for the H, which is apparented to the VH II type, and the L chains, which belong to the V kappa I subgroup. VH sequences are identical up to residue 43 and contain the common sequences (residues 1 to 32) defined for the H chains derived from the DBA/2 IgM anti-GAT monoclonal antibodies. Light chains are also remarkably conserved, a rather unusual situation for kappa-chains. The fifth antibody that expresses only part of the public idiotypic determinants contains very distinctive H and L chains. Its heavy chains are close to the VH I subgroup, whereas its kappa-chains permit definition of a new V kappa subgroup. The repertoire appears to be highly conserved between BALB/c and DBA/2 mice, and does not seem larger in IgG than in IgM antibodies. This latter observation does not speak in favor of a switch-linked amplification of diversity.

Amino Acid Sequence↗
PubMed1981

Organization of the diversity--joining region in rabbit immunoglobulin heavy chains as revealed by cleavage of a specific methionine residue in a100 allotype.

Three anti-micrococcus antibodies of restricted heterogeneity have been isolated from the antisera of homozygous a100/a100 rabbits. Heavy chains contained an unusual methionine residue at position 87 that may correlate with the a100 specificity. From this position, the sequence of a stretch of 35-50 amino acid residues was determined, permitting the definition of variable (V), diversity(D), and joining (J) segments in rabbit Ig heavy chains, with their most probable boundaries. Rabbit D regions so defined appear to be highly variable, both in sequence and in length, which varies, in the heavy chains analyzed, between 6 and 11 residues. The J regions are highly homologous to the mouse J2 segment.

Amino Acid Sequence↗
PubMed1976

Deletion of hinge region of human myeloma IgG1 molecule (protein LEC) associated with nonexpression of G1m (3) and Km (1, 2) allotypes. A possible genetic explanation at the DNA level.

In this paper we report the structural basis for the nonexpression of G1m(3) and Km (1,2) allotypes in an IgG1 (kappa) human myeloma protein (protein LEC). Heavy and light chains spontaneously dissociate in sodium dodecyl sulfate polyacrylamide gels. Light chains appear to be covalently S-S bonded. Analysis of cysteine-containing peptides shows that the heavy chain of the IgG protein LEC has a deletion of residues 216-230, thus encompassing the entire hinge region. An arginine residue, characteristic of the G1m(3) marker is present at position 214. An alanine at position 153 and a leucine at position 191 of the light chain, characteristic of the Km (1, 2) allotypes, are present. It is likely that the double Km and Gm lack of expression is the result of the deletion. The genetic implications of the sequence of this protein are discussed.

Amino Acid Sequence↗
PubMed1975

Determination of the primary structure of a mouse IgG2a immunoglobulin. Amino-acid sequence of the H4 cyanogen-bromide fragment.

The complete amino acid sequence of CNBr fragment H4 of the murine immunoglobulin MOPC 173 (IgG2a,chi) has been determined, thus completing the sequence determination of the entire heavy chain. The H4 fragment contains 150 residues, and extends from residue 105 to residue 254 of the heavy chain, which appears thus to be composed of 447 amino acids residues. This fragment contains the end of the V region, the switch peptide, the CH1 domain, the hinge region and the beginning of CH2. Sequence comparisons suggest that the CH1 domain is highly conserved in evolution, and allows the definition of two additional isotypic-specific regions.

Amino Acid Sequence↗
PubMed1975

Determination of the primary structure of a mouse IgG2a immunoglobulin. Amino-acid sequence of the light chain.

The amino acid sequence of the light chain of the mouse monoclonal MOPC 173 immunoglobulin molecule (IgG2a,chi) is presented. This kappa chain contains 214 residues. Comparisons of this sequence with murine kappa chains already published by other workers bring a confirmation of the large size of the murine V chi chain pool. A complete identity was found with the constant region of the light chain of MOPC 21 from residue 97 to residue 214.

Amino Acid Sequence↗