scholarly journals Synthetic peptides corresponding to third hypervariable region of human monoclonal IgM rheumatoid factor heavy chains define an immunodominant idiotype.

1985 ◽  
Vol 162 (2) ◽  
pp. 756-761 ◽  
Author(s):  
R D Goldfien ◽  
P P Chen ◽  
S Fong ◽  
D A Carson
Keyword(s):  
Rheumatoid Factor ◽  
Heavy Chain ◽  
Synthetic Peptides ◽  
Gene Segment ◽  
Hypervariable Region ◽  
Heavy Chains ◽  
The Third ◽  
Specific Induction ◽  
Antiidiotypic Antibodies ◽  
Complementarity Determining Regions

Synthetic peptides corresponding to eight individual heavy chain complementarity-determining regions (CDR) of three human monoclonal IgM anti-IgG (rheumatoid factor [RF]) paraproteins elicited rabbit antibodies with markedly different properties. All antisera recognized the immunizing peptide, and several reacted with the isolated IgM heavy chain on immunoblots. However, only the antisera against peptides representing the third CDR bound consistently and specifically to the intact IgM-RF molecule. These data indicate that the third CDR of human mu chains comprises an immunodominant idiotype, and suggest that the D gene segment may be especially important in creating idiotypic diversity. Synthetic peptides corresponding to the third heavy chain CDR of human paraproteins may be clinically useful for the specific induction of antiidiotypic antibodies.

scholarly journals Characterization of human rheumatoid factors with seven antiidiotypes induced by synthetic hypervariable region peptides.

1985 ◽  
Vol 162 (2) ◽  
pp. 487-500 ◽  
Author(s):  
P P Chen ◽  
F Gõni ◽  
R A Houghten ◽  
S Fong ◽  
R Goldfien ◽  
...  
Keyword(s):  
Rheumatoid Factor ◽  
Synthetic Peptides ◽  
Hypervariable Region ◽  
Rheumatoid Factors ◽  
Immunoblot Assay ◽  
The Third ◽  
Antiidiotypic Antibodies ◽  
Vh Genes ◽  
Complementarity Determining Regions

Recently, we have used synthetic peptides corresponding to the complementarity-determining regions (CDR) of Ig molecules to induce antiidiotypic antisera. Peptide PSH3, representing the third CDR of the IgM rheumatoid factor (RF) Sie heavy (H) chain, induced a private antiidiotype that reacted with only one out of five IgM-RF. Peptide PSL2, corresponding to the second CDR of Sie light (L) chain, induced an antibody against a crossreactive idiotype (CRI), expressed by 10 out of 12 human IgM-RF analyzed. Herein, we report that five additional antiidiotypic antibodies were generated by immunization with synthetic peptides identical to the third L chain CDR of IgM-RF Sie (PSL3), the second and third H chain CDR of IgM-RF Wol, and the second and third CDR of IgM-RF Pom. As analyzed by immunoblot assay, both anti-PSL3 and anti-PSL2 reacted with the majority of 16 IgM-RF. In contrast, all five antiidiotypes induced by the H chain peptides reacted only with the parent proteins, except anti-PSH3, which reacted weakly with one additional RF. These results suggest that one (or very few) VL gene(s), but a larger number of VH genes, are used to encode IgM-RF autoantibodies.

The diversity of rearranged immunoglobulin heavy chain variable region genes in peripheral blood B cells of preterm infants is restricted by short third complementarity-determining regions but not by limited gene segment usage

Blood ◽  
2001 ◽  
Vol 97 (5) ◽  
pp. 1511-1513 ◽  
Author(s):  
Michael Zemlin ◽  
Karl Bauer ◽  
Michael Hummel ◽  
Sabine Pfeiffer ◽  
Simone Devers ◽  
...  
Keyword(s):  
B Cells ◽  
Preterm Infants ◽  
Heavy Chain ◽  
Peripheral Blood ◽  
Gene Segment ◽  
Immunoglobulin Heavy Chain ◽  
Term Infants ◽  
Extremely Preterm ◽  
Extremely Preterm Infants ◽  
Complementarity Determining Regions

The immunoglobulin diversity is restricted in fetal liver B cells. This study examined whether peripheral blood B cells of extremely preterm infants show similar restrictions (overrepresentation of some gene segments, short third complementarity-determining regions [CDR3]). DNA of rearranged immunoglobulin heavy chain genes was amplified by polymerase chain reaction, cloned, and sequenced. A total of 417 sequences were analyzed from 6 preterm infants (25-28 weeks of gestation), 6 term infants, and 6 adults. Gene segments from the entire VHand DH gene locus were rearranged in preterm infants, even though the DH7-27 segment was overrepresented (17% of rearrangements) compared to term infants (7%) and adults (2%). CDR3 was shorter in preterm infants (40 ± 10 nucleotides) than in term infants (44 ± 12) and adults (48 ± 14) (P < .001) due to shorter N regions. Somatic mutations were exclusively found in term neonates and adults (mutational frequency 0.8% and 1.8%). We conclude that preterm infants have no limitations in gene segment usage, whereas the diversity of CDR3 is restricted throughout gestation.

An Atypical Human Immunoglobulin G with Deletions in both Heavy and Light Chains. Studies of the Conformation and the In Vitro Recombination of the Isolated Subunits

1974 ◽  
Vol 52 (7) ◽  
pp. 610-619 ◽  
Author(s):  
M. E. Percy ◽  
K. J. Dorrington
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
Hypervariable Region ◽  
Light Chains ◽  
Constant Region ◽  
Myeloma Protein ◽  
Variable Regions ◽  
Heavy Chains ◽  
Amino Terminal

Both the light and gamma (heavy) chains of IgG(Sac) contain extensive deletions in their variable regions. The deletion in the light chain is internal (residues 18–88), whereas the deletion in the heavy chain is amino-terminal (residues 1–102). The hypervariable region just preceding the beginning of the constant region in other heavy chains (residues 103–115) is amino-terminal in heavy chain(Sac). In 4 mM acetate, pH 5.4, heavy chain(Sac) is dimeric like normal gamma chains, whereas light chain(Sac) is monomeric. Isolated light and heavy chains of IgG(Sac) recombine in vitro with each other and also with the heavy and light chains from a typical human IgG1-K myeloma protein, but not in a fashion entirely typical of other human gamma and light chains. These studies support the concept that non-covalent forces between the variable regions of the light and heavy chains are important in the assembly of the immunoglobulin molecule; and in view of the weak interaction between the constant region of light chain and heavy chain observed previously, our data suggest that there are points of contact between the hypervariable region of the gamma chain (residues 103–115) and the variable region of the light chain.

scholarly journals Structural characterization of antiidiotypic antibodies. Evidence that Ab2s are derived from the germline differently than Ab1s.

1989 ◽  
Vol 169 (2) ◽  
pp. 519-533 ◽  
Author(s):  
K Meek ◽  
C Hasemann ◽  
B Pollok ◽  
S S Alkan ◽  
M Brait ◽  
...  
Keyword(s):  
Hypervariable Region ◽  
Variable Region ◽  
Good Evidence ◽  
Foreign Protein ◽  
Germline Gene ◽  
Region Gene ◽  
The Third ◽  
Antiidiotypic Antibodies ◽  
V Region

We have found that syngeneic Ab2s in the antiarsonate system are serologically and structurally similar to one another. In contrast, the allogeneic Ab2 response is heterogeneous and derives from a large number of unrelated germline gene segments. The Ab2 response of the BALB/c strain to polyclonal A/J Ars A molecules can probably best be compared with a response to a foreign protein and might have been predicted in a strain that completely lacks the H chain V region gene from which the Ab1 derives. Partial variable region sequences of Ab2s from three other systems in addition to previously reported Ab2 structures indicates that this difference in allogeneic vs. syngeneic Ab2s may be a general phenomena. These data support Jerne's hypothesis of complementary V region genes existing in the germline. However, there is good evidence that these antiidiotypic antibodies are not derived directly from the germline, as somatic processes most likely play an important role in their generation. The D segments of Ab2s in the arsonate system as well as in other systems, are novel in structure and cannot easily be explained by previously described germline D segments. D-D fusion may play a role in the generation of the third hypervariable region in these antibodies.

scholarly journals A discontinuous factor H binding site in the third component of complement as delineated by synthetic peptides.

1988 ◽  
Vol 263 (24) ◽  
pp. 12147-12150 ◽  
Author(s):  
J D Lambris ◽  
D Avila ◽  
J D Becherer ◽  
H J Müller-Eberhard
Keyword(s):  
Binding Site ◽  
Synthetic Peptides ◽  
Factor H ◽  
The Third ◽  
Third Component Of Complement

Acidic Region Residues 1680–1684 in the A3 Domain of Factor VIII Contain a Thrombin-Interactive Site Responsible for Proteolytic Cleavage at Arg1689

2021 ◽  
Author(s):  
Yuto Nakajima ◽  
Hiroaki Minami ◽  
Keiji Nogami
Keyword(s):  
Surface Plasmon Resonance ◽  
Factor Viii ◽  
Heavy Chain ◽  
Synthetic Peptides ◽  
C2 Domain ◽  
Wild Type ◽  
Cross Link ◽  
Domain Specific ◽  
Cofactor Activity ◽  
Acidic Region

AbstractFactor VIII (FVIII) is activated by thrombin-catalyzed cleavage at Arg372, Arg740, and Arg1689. Our previous studies suggested that thrombin interacted with the FVIII C2 domain specific for cleavage at Arg1689. An alternative report demonstrated, however, that a recombinant (r)FVIII mutant lacking the C2 domain retained >50% cofactor activity, indicating the presence of other thrombin-interactive site(s) associated with cleavage at Arg1689. We have focused, therefore, on the A3 acidic region of FVIII, similar to the hirugen sequence specific for thrombin interaction (54–65 residues). Two synthetic peptides, spanning residues 1659–1669 with sulfated Tyr1664 and residues 1675–1685 with sulfated Try1680, inhibited thrombin-catalyzed FVIII activation and cleavage at Arg1689. Treatment with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide to cross-link thrombin with either peptide showed possible contributions of both 1664–1666 and 1683–1684 residues for thrombin interaction. Thrombin-catalyzed activation and cleavage at Arg1689 in the alanine-substituted rFVIII mutants within 1663–1666 residues were similar to those of wild type (WT). Similar studies of 1680–1684 residues, however, demonstrated that activation and cleavage by thrombin of the FVIII mutant with Y1680A or D1683A/E1684A, in particular, were severely or moderately reduced to 20 to 30% or 60 to 70% of WT, respectively. Surface plasmon resonance-based analysis revealed that thrombin interacted with both Y1680A and D1683A/E1684A mutants with approximately sixfold weaker affinities of WT. Cleavage at Arg1689 in the isolated light-chain fragments from both mutants was similarly depressed, independently of the heavy-chain subunit. In conclusion, the 1680–1684 residues containing sulfated Tyr1680 in the A3 acidic region also contribute to a thrombin-interactive site responsible for FVIII activation through cleavage at Arg1689.

Synthetic idiotypes: The third hypervariable region of murine anti-dextran antibodies

Cell ◽  
1983 ◽  
Vol 35 (3) ◽  
pp. 859-863 ◽  
Author(s):  
S. McMillan ◽  
M.V. Seiden ◽  
R.A. Houghten ◽  
B. Clevinger ◽  
J.M. Davie ◽  
...  
Keyword(s):  
Hypervariable Region ◽  
The Third

scholarly journals A Two-Step Approach for the Design and Generation of Nanobodies

2018 ◽  
Vol 19 (11) ◽  
pp. 3444 ◽  
Author(s):  
Hanna Wagner ◽  
Sarah Wehrle ◽  
Etienne Weiss ◽  
Marco Cavallari ◽  
Wilfried Weber
Keyword(s):  
Heavy Chain ◽  
Animal Experiments ◽  
Affinity Maturation ◽  
Phage Library ◽  
Second Step ◽  
Selection Procedures ◽  
Variable Domains ◽  
Phage Libraries ◽  
Complementarity Determining Regions ◽  
Conventional Antibody

Nanobodies, the smallest possible antibody format, have become of considerable interest for biotechnological and immunotherapeutic applications. They show excellent robustness, are non-immunogenic in humans, and can easily be engineered and produced in prokaryotic hosts. Traditionally, nanobodies are selected from camelid immune libraries involving the maintenance and treatment of animals. Recent advances have involved the generation of nanobodies from naïve or synthetic libraries. However, such approaches demand large library sizes and sophisticated selection procedures. Here, we propose an alternative, two-step approach for the design and generation of nanobodies. In a first step, complementarity-determining regions (CDRs) are grafted from conventional antibody formats onto nanobody frameworks, generating weak antigen binders. In a second step, the weak binders serve as templates to design focused synthetic phage libraries for affinity maturation. We validated this approach by grafting toxin- and hapten-specific CDRs onto frameworks derived from variable domains of camelid heavy-chain-only antibodies (VHH). We then affinity matured the hapten binder via panning of a synthetic phage library. We suggest that this strategy can complement existing immune, naïve, and synthetic library based methods, requiring neither animal experiments, nor large libraries, nor sophisticated selection protocols.

scholarly journals Chlamydomonas WDR92 in association with R2TP-like complex and multiple DNAAFs to regulate ciliary dynein preassembly

2018 ◽  
Vol 11 (9) ◽  
pp. 770-780 ◽  
Author(s):  
Guang Liu ◽  
Limei Wang ◽  
Junmin Pan
Keyword(s):  
Mass Spectrometry ◽  
Insertional Mutagenesis ◽  
The Other ◽  
Cytoplasmic Protein ◽  
Flagellar Motility ◽  
Heavy Chains ◽  
The Third ◽  
Axonemal Dynein ◽  
Dynein Arms ◽  
Dna Insertional Mutagenesis

Abstract The motility of cilia or eukaryotic flagella is powered by the axonemal dyneins, which are preassembled in the cytoplasm by proteins termed dynein arm assembly factors (DNAAFs) before being transported to and assembled on the ciliary axoneme. Here, we characterize the function of WDR92 in Chlamydomonas. Loss of WDR92, a cytoplasmic protein, in a mutant wdr92 generated by DNA insertional mutagenesis resulted in aflagellate cells or cells with stumpy or short flagella, disappearance of axonemal dynein arms, and diminishment of dynein arm heavy chains in the cytoplasm, suggesting that WDR92 is a DNAAF. Immunoprecipitation of WDR92 followed by mass spectrometry identified inner dynein arm heavy chains and multiple DNAAFs including RuvBL1, RPAP3, MOT48, ODA7, and DYX1C. The PIH1 domain-containing protein MOT48 formed a R2TP-like complex with RuvBL1/2 and RPAP3, while PF13, another PIH1 domain-containing protein with function in dynein preassembly, did not. Interestingly, the third PIH1 domain-containing protein TWI1 was not related to flagellar motility. WDR92 physically interacted with the R2TP-like complex and the other identified DNNAFs. Our data suggest that WDR92 functions in association with the HSP90 co-chaperone R2TP-like complex as well as linking other DNAAFs in dynein preassembly.

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