scholarly journals Goat immunoglobin G. Peptide chains and terminal residues

1969 ◽  
Vol 115 (3) ◽  
pp. 371-375 ◽  
Author(s):  
D. Givol ◽  
E. Hurwitz
Keyword(s):  
Carboxylic Acid ◽  
Heavy Chain ◽  
Light Chain ◽  
Immunoglobulin G ◽  
Light Chains ◽  
Terminal Sequence ◽  
Terminal Residue ◽  
Heavy Chains ◽  
Molecular Weights ◽  
Immunoglobin G

Goat immunoglobulin G (IgG) was isolated and characterized. The molecular weights of the IgG and its heavy chains and light chains were found to be 144000, 53600 and 23000 respectively. The light chain corresponds to human L type as was shown by the absence of C-terminal S-carboxymethylcysteine and its high content of N-terminal pyrrolid-2-one-5-carboxylic acid (PCA). The major C-terminal residue of the light chain was serine and the major N-terminal dipeptide was PCA-Ala (0·6mole/mole). The major C-terminal residue of the heavy chain was glycine and the N-terminal sequence of the heavy chain is PCA-Val-Gln. This tripeptide was obtained in a 70% yield.

scholarly journals An Atypical Immunoglobulin

Blood ◽  
1968 ◽  
Vol 32 (2) ◽  
pp. 189-204 ◽  
Author(s):  
A. F. LEWIS ◽  
D. E. BERGSAGEL ◽  
A. BRUCE-ROBERTSON ◽  
R. K. SCHACHTER ◽  
G. E. CONNELL
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
Gel Filtration ◽  
Sedimentation Equilibrium ◽  
Light Chains ◽  
Equilibrium Studies ◽  
Heavy Chains ◽  
Molecular Weights ◽  
Plasma Cell Neoplasm ◽  
Ig G

Abstract A protein of the Ig G family has been isolated from the serum of a patient with a tentative diagnosis of a plasma cell neoplasm. The protein has a lower sedimentation constant (5.4) and a lower molecular weight (125,000) than normal immunoglobulins of the G family. The protein has heavy-chain determinants of type G and light-chain determinants of the κ-type. Heavy and light chains have been prepared by reductive cleavage followed by gel filtration. The heavy-chain preparation is homogeneous in starch gels in acidic buffer containing urea but has a faster mobility than normal Ig G heavy chains. The light-chain preparation is resolved into two components in electrophoresis, and both have slower mobility than normal Ig G light chains. The heavy- and light-chain preparations cross react with normal Ig G heavy and light chains in immunodiffusion analysis. Sedimentation equilibrium studies suggest that both the heavy and light chains have lower molecular weights than their normal counterparts.

scholarly journals A blocked N-terminal residue in the light chain of rabbit immunoglobulin G

1968 ◽  
Vol 107 (4) ◽  
pp. 449-453 ◽  
Author(s):  
E. Rüde ◽  
D. Givol
Keyword(s):  
Carboxylic Acid ◽  
Light Chain ◽  
Immunoglobulin G ◽  
Terminal Sequence ◽  
Terminal Residue ◽  
Rabbit Immunoglobulin

The light chain of rabbit immunoglobulin G was shown to contain 15–20% blocked N-terminal residue. The blocked residue is pyrrolid-2-one-5-carboxylic acid, and most of the chains that contain this residue have the N-terminal sequence pyrrolid-2-one-5-carbonyl-valine.

scholarly journals Intrachain disulphide bridges in immunoglobulin G heavy chains. The Fc fragment

1968 ◽  
Vol 106 (1) ◽  
pp. 15-21 ◽  
Author(s):  
B. Frangione ◽  
C. Milstein ◽  
Edward C. Franklin
Keyword(s):  
Heavy Chain ◽  
Immunoglobulin G ◽  
The Other ◽  
Light Chains ◽  
Fc Fragment ◽  
Heavy Chains ◽  
Human Immunoglobulins ◽  
Other Hand ◽  
Terminal Loop ◽  
Heavy Chain Disease

The disulphide bridges of the Fc fragment (C-terminal half of the heavy chain) have been studied in several human immunoglobulins, containing heavy chains of different antigenic types (γ1, γ2, γ3 and γ4), and in heavy-chain-disease proteins. Two intrachain disulphide bridges were found to be present. The sequences appear to be identical in the Fc fragments of two types of chain studied (γ1 and γ3), and very similar to corresponding sequences of the Fc fragment in rabbit. These results suggest that the C-terminal half of the heavy chains is covalently folded (in a similar fashion to the light chains) with a C-terminal loop and an N-terminal loop. The similarity is emphasized by comparison of the sequence and location of the disulphide-bridged peptides of the C-terminal loop of heavy and light chains. The N-terminal loop, on the other hand, appears to be very different in Fc fragments and light chains. The C-terminal loop is the only one present in the F′c fragment.

Inhibitory ADAMTS13 Monoclonal Autoantibodies Cloned from TTP Patients Show Restricted Gene Usage, Inhibit Murine ADAMTS13, and Are Blocked by Rabbit Anti-Idiotypic Antibodies.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 92-92 ◽  
Author(s):  
Don Siegel ◽  
Eric Ostertag
Keyword(s):  
Immune Response ◽  
Phage Display ◽  
Heavy Chain ◽  
Light Chain ◽  
Light Chains ◽  
Chain Gene ◽  
Human Mabs ◽  
Heavy Chains ◽  
Phage Display Libraries ◽  
Pathogenic Antibodies

Abstract Thrombotic thrombocytopenic purpura (TTP) is a potentially fatal disorder often associated with autoantibody inhibition of ADAMTS13, a VWF-cleaving protease. Autoantibodies decrease ADAMTS13 activity resulting in accumulation of “unusually” large VWF multimers that mediate platelet thrombosis. To better understand the role autoantibodies play in disease pathogenesis, as well as to develop more specific methods for diagnosis and therapy, it is necessary to characterize pathogenic antibodies on a molecular level, something not possible through analysis of polyclonal patient antisera. The ability to clone large repertoires of patient monoclonal autoantibodies (mAbs) using phage display offers a unique opportunity to address this issue. Three patient (Pt) antibody phage display libraries were created from either splenocytes (Pt1) or peripheral blood lymphocytes (Pt2, Pt3) of individuals with acquired TTP. ADAMTS13-specific mAbs were isolated by panning against recombinant ADAMTS13. Unique clones were identified by DNA sequencing, and their ability to interact with ADAMTS13 was characterized. After antigen selection of Pt1 library, 56 mAbs were randomly-selected from panning rounds 2 through 4 and 68% were found to comprise heavy chains encoded by VH1-69 paired with a VL3 family lambda light chain (3h or 3m). The remaining mAbs comprised heavy chains from the VH1, 3, or 4 families usually paired with kappa light chains. For Pt2 and Pt3 libraries, there was an identical pattern of genetic restriction in immune response to ADAMTS13, i.e. 16 of 24 mAbs (Pt2) and 27 of 27 mAbs (Pt3) were encoded by VH1-69 heavy chains and VL3 family lambda light chains. Though nearly all mAbs were unique, common CDR3 regions among some of the mAbs provided evidence of B-cell clonal expansion and somatic mutation. Though all mAbs bound to ADAMTS13 irrespective of genetic origin, mAbs comprising a VH1-69 heavy chain paired with a VL3 light chain inhibited ADAMTS13 using the FRET-VW73 assay while mAbs comprising a VH1-69 paired with a kappa light chain or comprising non-VH1-69 heavy chains did not inhibit ADAMTS13, with only two exceptions. MAb binding to ADAMTS13 was blocked by preincubation with normal human or murine plasma, but much less so by plasma from TTP patients or ADAMTS13 knockout mice suggesting crossreactivity with mouse ADAMTS13. Certain human mAbs inhibited cleavage of FRET-VWF73 by mouse ADAMTS13 and also inhibited ADAMTS13 in vivo after injection into the internal jugular vein of mice. Rabbit anti-idiotypic antibodies raised against mAb 416, a prototypical VH1-69-encoded mAb, blocked 416’s ability to inhibit human ADAMTS13. Taken together, the cloning and analyses of a large cohort of ADAMTS13 inhibitory autoantibodies derived from 3 unrelated individuals with acquired TTP revealed a genetically restricted immune response. This feature, if common among TTP patients, offers a potential therapeutic target for treatment of TTP, e.g. selective deletion of B-cells utilizing the VH1-69 heavy chain gene. Furthermore, crossreactivity of some human mAbs with murine ADAMTS13 provides a mouse model of acquired ADAMTS13 deficiency that may prove useful for determining the role of autoantibodies in the pathogenesis of TTP, particularly in the context of additional factors (e.g. environmental) that may be required to trigger the disease. Finally, anti-idiotypic mAbs, currently being cloned from rabbit phage display libraries, may help identify pathogenic antibodies in patient plasma and/or lead to novel therapeutic approaches.

scholarly journals The formation of pyrrolid-2-one-5-carboxylic acid at the N-terminus of immunoglobulin G heavy chain

1972 ◽  
Vol 128 (5) ◽  
pp. 1221-1227 ◽  
Author(s):  
D. I. Stott ◽  
A. J. Munro
Keyword(s):  
Protein Synthesis ◽  
Carboxylic Acid ◽  
Heavy Chain ◽  
Immunoglobulin G ◽  
Acid Group ◽  
Eukaryotic Cells ◽  
Double Labelling ◽  
Carboxylic Acid Group ◽  
Heavy Chains ◽  
N Terminus

We propose that pyrrolid-2-one-5-carboxyl-tRNA is not involved in the initiation of protein synthesis in eukaryotic cells and that the N-terminal pyrrolid-2-one-5-carboxylic acid group of an IgG (immunoglobulin G) (that secreted by the mouse plasmacytoma Adj PC5) is formed by the enzymic cyclization of the N-terminal glutamine of the heavy chain of the completed IgG molecule and that the cyclization takes place inside the cell. We base these conclusions on the following evidence. (1) Pyrrolidonecarboxyl-tRNA was not found in incorporation experiments with rat liver preparations and [U-14C]-pyrrolidonecarboxylic acid, glutamic acid and glutamine, even though an incorporation extent of less than 2% of the total products could have been detected. (2) Double-labelling experiments showed that less than 8% of the nascent peptides of heavy chains (those obtained by precipitation by the antibody to Fc fragment) began with pyrrolidonecarboxylic acid. (3) Further double-labelling experiments showed that 60–66% of the heavy chains of the completed intracellular IgG molecule began with pyrrolidonecarboxylic acid after both 1 and 5h of labelling. (4) The IgG, after secretion by plasmacytoma Adj PC5, was found to have the sequence [unk]Glu- Val-Gln-Leu- at the N-termini of the heavy chains.

Novel functions of clathrin light chains: clathrin heavy chain trimerization is defective in light chain-deficient yeast

1997 ◽  
Vol 110 (7) ◽  
pp. 899-910 ◽  
Author(s):  
K.M. Huang ◽  
L. Gullberg ◽  
K.K. Nelson ◽  
C.J. Stefan ◽  
K. Blumer ◽  
...  
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
State Level ◽  
Light Chains ◽  
Chain Gene ◽  
Light Chain Gene ◽  
Heavy Chains ◽  
Clathrin Heavy Chain ◽  
Multicopy Suppressors ◽  
Delta Cells

Clathrin is a major coat protein involved in sorting and retention of proteins at the late Golgi and in endocytosis from the cell surface. The clathrin triskelion contains three heavy chains, which provide the structural backbone of the clathrin lattice and three light chains, which are thought to regulate the formation or disassembly of clathrin coats. To better understand the function of the clathrin light chain, we characterized yeast strains carrying a disruption of the clathrin light chain gene (CLC1). Light chain-deficient cells showed phenotypes similar to those displayed by yeast that have a disruption in the clathrin heavy chain gene (CHC1). In clc1-delta cells, the steady state level of the clathrin heavy chain was reduced to 20%-25% of wild-type levels and most of the heavy chain was not trimerized. If CHC1 was overexpressed in clc1-delta cells, heavy chain trimers were detected and several clc1-delta phenotypes were partially rescued. These results indicate that the light chain is important for heavy chain trimerization and the heavy chain still has some function in the absence of the light chain. In yeast, deletion of CHC1 is lethal in strains carrying the scd1-i allele, while strains carrying the scd1-v allele can survive without the heavy chain. In previous studies we isolated several multicopy suppressors of inviability of chc1-delta scd1-i cells. Surprisingly, one of these suppressors, SCD4, is identical to CLC1. Overexpression of CLC1 in viable chc1-delta scd1-v strains rescued some but not all of the phenotypes displayed by these cells. In the absence of the heavy chain, the light chain was not found in a high molecular mass complex, but still associated with membranes. These results suggest that the light chain can function independently of the clathrin heavy chain in yeast.

scholarly journals Study of Basic Local Alignment Search Tool (BLAST) and Multiple Sequence Alignment (Clustal- X) of Monoclonal mice/human antibodies

2021 ◽  
Author(s):  
IVAN VITO FERRARI ◽  
Paolo PATRIZIO
Keyword(s):  
Amino Acids ◽  
Monoclonal Antibodies ◽  
Sequence Alignment ◽  
Heavy Chain ◽  
Light Chain ◽  
Light Chains ◽  
Local Alignment ◽  
Multiple Sequence ◽  
Heavy Chains ◽  
Search Tool

In this work, we have focused on the study of the Basic Local Alignment Search Tool (BLAST) and Multiple Sequence Alignment (Clustal- X) of different monoclonal mice antibodies to understand better the multiple alignments of sequences. Our strategy was to compare the light chains of multiple monoclonal antibodies to each other, calculating their identity percentage and in which amino acid portion. (See below figure 2) Subsequently, the same survey of heavy chains was carried out with the same methodology. (See below figure 3) Finally, sequence alignment between the light chain of one antibody and the heavy chain of another antibody was studied to understand what happens if chains are exchanged between antibodies. (See below figure 4) From our results of BLAST estimation alignment, we have reported that the Light Chains (Ls) of Monoclonal Antibodies in Comparison have a sequence Homology of about 60-80% and they have a part identical in sequence zone in range 100-210 residues amino acids, except ID PDB 4ISV, which it turns out to have a 40% lower homology than the others antibodies. As far as, the heavy chains (Hs) of Monoclonal Antibodies are concerned, however they tend to have a less homology of sequences, compared to lights chains consideration, equal to 60%-70% and they have an identical part in the sequence zone between 150-210 residues amino acids; with the exception of ID PDB 3I9G-3W9D antibodies that have an equal homology at 50%. ( See supporting part) Summing up: about 70-80% identity among 2 light chains of 2 antibodies, 60-70% identity between 2 heavy chains of 2 antibodies, 30% identity between the two chains of a antibody and 30% if you compare the light chain of one antibody with the heavy chain of another antibody.

Evidence for the Significant Role of Immunoglobulin Light Chains in Antigen Recognition and Selection in Chronic Lymphocytic Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 780-780
Author(s):  
Anastasia Hadzidimitriou ◽  
Nikos Darzentas ◽  
Fiona Murray ◽  
Tanja Smilevska ◽  
Eleni Arvaniti ◽  
...  
Keyword(s):  
Amino Acid ◽  
Chronic Lymphocytic Leukemia ◽  
Heavy Chain ◽  
Light Chain ◽  
Antigen Recognition ◽  
Lymphocytic Leukemia ◽  
Light Chains ◽  
Light Chain Gene ◽  
Heavy Chains ◽  
Gene Usage

Abstract The chronic lymphocytic leukemia (CLL) immunoglobulin (IG) heavy chain repertoire is known to display biased immunoglobulin variable heavy-chain (IGHV) gene usage, remarkable complementarity determining region 3 (HCDR3) stereotypy as well as distinctive somatic hypermutation (SHM) patterns, at least for subsets of cases. Our aim in the present study was to similarly investigate the IG light chain (LC) genes in terms of mutation frequency and targeting and CDR3 stereotypy to elucidate if the LC may play a significant complementary role in antigen recognition in CLL. We thus examined SHM patterns and secondary rearrangements of the IG LC gene loci in a total of 612 IGKV-J and 279 IGLV-J rearrangements from 725 patients with CLL. Firstly, we observed a highly restricted light chain gene usage in the vast majority of CLL cases with stereotyped HCDR3s. In particular, stereotyped IGHV3-21 CLL cases were characterized by a strikingly biased expression of lambda light chains utilizing the IGLV3-21 gene (36/37 cases of subset#2), whereas all 15 subset #4 cases with stereotyped IGHV4-34 IGs carried an IGKV2-30 rearrangement. In addition, subset-biased light chain CDR3 motifs were identified in groups of sequences utilizing the same IGKV or IGLV gene. For example, all 30 IGKV1-39/1D-39 light chains of subset#1 (using stereotyped IGHV1/5/7 genes) carried notably long KCDR3s (10–11 amino acids) generated by significant N region addition and characterized by the frequent introduction of a junctional proline (26/30 cases). Important differences regarding mutational load were observed in groups of sequences utilizing the same IGKV or IGLV gene and/or belonging to subsets with stereotyped B cell receptors (BCRs). In fact, significant differences were observed with regard to mutational status among groups of sequences utilizing different alleles of certain IGK/LV genes (specifically the IGKV1-5, IGLV1-51 and IGLV3-21 genes). At cohort level, the SHM patterns were typical of a canonical SHM process. A clustering of R mutations in KCDR1 was evident for all IGKV subgroups with the notable exception of the IGKV2 subgroup, which exhibited preferential targeting to the KCDR2, especially in IGKV2-30 rearrangements of cases with stereotyped IGHV4-34/IGKV2-30 BCRs (subset#4). Recurrent amino acid changes at certain positions across the entire IGKV/IGLV sequence were observed at a high frequency (27–67% of cases) in a number of stereotyped subsets, especially those expressing the IGHV3-21/IGLV3-21 BCR (subset #2) and the IGHV4-34/IGKV2-30 BCR (subset #4). Comparison with CLL LC sequences carrying heterogeneous K/LCDR3s or non-CLL LC sequences revealed that these distinct amino acid changes are greatly under-represented in such groups and appear therefore to be “subset-biased”. Finally, a significant proportion of CLL cases (63 cases; 26 kappa- and 37 lambda-expressing) with monotypic LC expression were found to carry multiple potentially functional LC rearrangements. Of note, nineteen of these 63 cases (30%) belonged to subsets with stereotyped BCRs. This finding alludes to the possibility of secondary rearrangements most likely occurring in the context of (auto)antigen-driven receptor editing, particularly in the case of stereotyped subsets. In conclusion, SHM targeting in CLL LCs appears to be just as precise and, most likely, functionally driven as in heavy chains. Secondary LC gene rearrangements and subset-biased mutations in CLL LC genes are strong indications that LCs are crucial in shaping the specificity of leukemic BCRs, in association with defined heavy chains. Therefore, CLL is characterized not only by stereotyped HCDR3 and heavy chains but, rather, by stereotyped BCRs involving both chains, which create distinctive antigen binding grooves.

Studies on the amino acid sequence of heavy chains from rabbit immunoglobulin G

1966 ◽  
Vol 166 (1003) ◽  
pp. 159-175 ◽  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Heavy Chain ◽  
Immunoglobulin G ◽  
Light Chains ◽  
Antibody Specificity ◽  
Sequence Heterogeneity ◽  
Specific Antibodies ◽  
Heavy Chains ◽  
Insight Into

It is now generally agreed that the four-chain subunit structure of Immunoglobulins which was first proposed by Porter (1962), accurately represents the gross structure of immunoglobulin G (IgG) and specific antibodies (Fleischman, Porter & Press 1963; Edelman & Gally 1964; Marler, Nelson & Tanford 1964; Nelson et al . 1965). However, an understanding of the structural basis of antibody specificity requires greater insight into the amino acid sequence of the polypeptide chain components of specific antibodies. Isolated light chains from specific antibodies and inert IgG, show a considerable degree of electrophoretic heterogeneity (Edelman & Gally 1964; Cohen & Porter 1964; Poulik 1964). Tryptic peptide maps of light chains (Nelson et al . 1965) have suggested that this heterogeneity may be accounted for by differences in amino acid sequence. This view has received considerable support from the observation that Bence-Jones proteins, which may be regarded as light chains, vary significantly in amino acid sequence (Hilschman & Craig 1965; Milstein 1966; Titani, Whitley & Putman 1966). A similar but less well-defined sequence heterogeneity has been suggested to exist in the heavy chains of specific antibodies (Feinstein 1964). However, the Fc fragment of the heavy chains has been thought to possess a regular amino acid sequence which may be similar, if not identical, among all specific antibodies (Porter 1959; Nelson et al . 1965). This paper summarizes the results of studies on the amino acid sequence of heavy chains and that portion of heavy chain, Fc fragment, which is obtained on treatment of rabbit IgG with papain (Porter 1959). These studies were designed to determine how much of the amino acid sequence of heavy chain could be accounted for by a unique, regular amino acid sequence which was common to most, if not all, IgG antibodies. In addition, attempts were made to locate regions of heavy chains which varied in amino acid sequence. Although structural variants appear to occur among the heavy chains found in non-specific IgG, it would be desirable to know what portion of the heavy chain sequence is invariant among all antibodies. If antibody specificity results from sequence heterogeneity in light and heavy chains, then knowledge of the variant and invariant portions of these chains may provide insight into the nature of specific binding sites in anti-­bodies.

Specificity of antigenic recognition of antibody heavy chain

1966 ◽  
Vol 166 (1003) ◽  
pp. 176-187 ◽  
Keyword(s):  
Heavy Chain ◽  
Light Chain ◽  
Group Analysis ◽  
Binding Activity ◽  
Association Constants ◽  
Light Chains ◽  
Specific Information ◽  
Parent Molecule ◽  
Heavy Chains ◽  
Antibody Heavy Chain

The specificity of antigenic recognition of the component chains of purified dinitrophenyl and trinitrophenyl antibodies was examined. Heavy chains were rendered soluble at neutral pH, either by prior reaction of the parent antibodies with D,L-alanine N -carboxy anhydride, or by mixing heavy chain with light chain of non-specific IgG. The degree of homologous light chain contamination of these heavy chain preparations was found to be less than 2 %, either by immune precipitation, or by end-group analysis. Association constants of the heavy chains of both antibodies with several closely related haptenes were measured by fluorescence quenching. Heavy chains differentiated among these haptenes in the same manner as the parent antibodies, though considerable binding affinity was lost. When specific homologous light chains were added to the heavy chain preparations, association constants were increased, but without change in relative selectivity. Binding activity of light chains alone could not be measured. The heavy chain, then, appears to bear the specificity of its parent molecule. Whether or not homologous light chain contributes additional specific information with respect to antigenic recognition or simply plays a non-specific modulating role cannot be answered from these experiments.

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