Regulation of Antibody Activity by Pro- and Anti-oxidants via Modification of the Disulfide Bonds in Antibody Structure

The elasmobranch Mustelus canis has been shown to produce antibodies to Limulus hemocyanin. The serum of both normal and immunized M. canis contains immunoglobulins having sedimentation coefficients of approximately 7S and 17S. Antibody activity was found in the 17S immunoglobulin which may be dissociated to 7S components with concomitant loss of activity. Both 17S and 7S serum, immunoglobulins were antigenically identical. They consisted of light and heavy chains present in amounts comparable to those of higher vertebrates. Peptide maps indicated that the light chains had an entirely different primary structure than the heavy chains, but that the corresponding chains of 7S and 17S dogfish serum immunoglobulins were similar in primary structure. The heavy chains appeared to resemble the n chains of immunoglobulins of higher vertebrates in their starch gel electrophoretic behavior. It is suggested that the elasmobranch M. canis may have only one major class of immunoglobulins resembling that of macroglobulins (γM-immunoglobulins) seen in higher vertebrates. The results indicate that the multichain structure of antibodies is an ancient evolutionary development.

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Mechanistic principles of an ultra-long bovine CDR reveal strategies for antibody design

Nature Communications ◽

10.1038/s41467-021-27103-z ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Hristo L. Svilenov ◽

Julia Sacherl ◽

Ulrike Protzer ◽

Martin Zacharias ◽

Johannes Buchner

Keyword(s):

Disulfide Bonds ◽

High Efficiency ◽

De Novo ◽

Human Antibody ◽

Antigen Binding ◽

Antibody Structure ◽

Alpha Variant ◽

Complementarity Determining Regions ◽

Antibody Design ◽

Stalk Length

AbstractAntibodies bind antigens via flexible loops called complementarity-determining regions (CDRs). These are usually 6-20 residues long. However, some bovine antibodies have ultra-long CDRs comprising more than 50 residues organized in a stalk and a disulfide-rich knob. The design features of this structural unit and its influence on antibody stability remained enigmatic. Here, we show that the stalk length is critical for the folding and stability of antibodies with an ultra-long CDR and that the disulfide bonds in the knob do not contribute to stability; they are important for organizing the antigen-binding knob structure. The bovine ultra-long CDR can be integrated into human antibody scaffolds. Furthermore, mini-domains from de novo design can be reformatted as ultra-long CDRs to create unique antibody-based proteins neutralizing SARS-CoV-2 and the Alpha variant of concern with high efficiency. Our findings reveal basic design principles of antibody structure and open new avenues for protein engineering.

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CHARACTERISTICS OF AN IMMUNE SYSTEM COMMON TO CERTAIN EXTERNAL SECRETIONS

Journal of Experimental Medicine ◽

10.1084/jem.121.1.101 ◽

1965 ◽

Vol 121 (1) ◽

pp. 101-124 ◽

Cited By ~ 684

Author(s):

Thomas B. Tomasi ◽

Eng M. Tan ◽

Alan Solomon ◽

Robert A. Prendergast

Keyword(s):

Disulfide Bonds ◽

Defense Mechanisms ◽

Sedimentation Coefficient ◽

Antibody Activity ◽

Local Production ◽

Local Synthesis ◽

Distinctive Type ◽

Γ Globulins ◽

Polypeptide Chains ◽

Circulating Antibody

The γ1A present in saliva and colostrum exists largely in the form of higher polymers, the major component of which has a sedimentation coefficient of 11S. The 11S γ1A in these fluids differs from the polymers found in normal and myeloma sera both immunologically and by the fact that their sedimentation coefficients are unaffected by disulfide bond reduction in the absence of urea. However, like other γ-globulins the 11S γ1A molecules consist of multiple polypeptide chains linked by disulfide bonds. Local synthesis of γ1A in the salivary gland has been shown by fluorescent and autoradiographic studies, although the fraction of the total salivary γ1A which is derived from local production is uncertain. No evidence of transport of intravenously administered I131-labeled 7S γ1A from serum to saliva was obtained. Immunological specificity has been demonstrated in the salivary and colostral γ1A. Whether that portion of the γ1A which is immunologically specific is a piece incorporated during the local synthesis of γ1A in the gland or is added by the epithelial cell in the process of transport remains to be determined. Antibody activity (isohemagglutinins) have been demonstrated in saliva and colostrum and have been shown to be of the γ1A-type. In both of these fluids activity is associated primarily with γ1A-polymers of 11S and 18S sizes. There appears to be an immunological system which is characteristic of certain external secretions. Its properties including the local production of a distinctive type of antibody separate it from the "systemic" system responsible for the production of circulating antibody. This system may play a significant role in the body's defense mechanisms against allergens and microorganisms.

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PHYLOGENETIC ORIGINS OF ANTIBODY STRUCTURE

Journal of Experimental Medicine ◽

10.1084/jem.127.5.891 ◽

1968 ◽

Vol 127 (5) ◽

pp. 891-914 ◽

Cited By ~ 136

Author(s):

John J. Marchalonis ◽

Gerald M. Edelman

Keyword(s):

Molecular Weight ◽

Heavy Chain ◽

Disulfide Bonds ◽

Serum Proteins ◽

Gel Filtration ◽

Exchange Chromatography ◽

Antibody Activity ◽

Starch Gel Electrophoresis ◽

Antigenic Analysis ◽

Amino Terminal

The sea lamprey, Petromyzon marinus, has been found to produce specific antibodies after immunization with bacteriophage f2. Antibody activity is localized in 6.6S and 14S fractions of lamprey serum. The 6.6S antibodies were purified by a combination of zone electrophoresis, ion exchange chromatography, and gel filtration. Antigenic analysis of the 6.6S antibodies showed them to be free of other serum proteins and antigenically similar or identical to the 14S fraction. Evidence has been obtained which suggests that the 6.6S immunoglobulins consist of light components (molecular weight 25,000) and heavy components (molecular weight 70,000). In the immunoglobulin, these polypeptides appear to be linked via weak interactions but not by interchain disulfide bonds. Molecular weight analyses support the view that the chains can undergo concentration-dependent dissociation in aqueous solutions. Amino acid analyses showed that the compositions of the light and heavy components were similar and that aspartic acid or asparagine was the predominant amino terminal residue. Starch gel electrophoresis indicated that the subunits of lamprey antibodies are diffusely heterogeneous. The heavy chain mobility corresponded to that of µ-chains and resembled that of heavy chains of shark and sting ray immunoglobulins. In the course of the fractionation a 46S natural hemagglutinin composed of lower molecular weight subunits was isolated. This hemagglutinin did not resemble the lamprey immunoglobulin although it had a similar zone electrophoretic mobility in the ß-region. These studies are consistent with the hypothesis that µ-chains were the earliest of the heavy chain classes to emerge and further support the view that the multichain structure of immunoglobulins is a fundamental feature of antibody molecules.

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Cryo-electron microscopy of two-dimensional crystals of reduced type B botulinum neurotoxin

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100123052 ◽

1992 ◽

Vol 50 (1) ◽

pp. 530-531

Author(s):

P. F. Flicker ◽

V.S. Kulkarni ◽

J. P. Robinson ◽

G. Stubbs ◽

B. R. DasGupta

Keyword(s):

Disulfide Bonds ◽

Clostridium Botulinum ◽

Structural Changes ◽

Two Dimensional ◽

Type B ◽

Single Chain ◽

Muscle Paralysis ◽

Cryo Electron Microscopy ◽

Disulfide Linkages ◽

Intracellular Action

Botulinum toxin is a potent neurotoxin produced by Clostridium botulinum. The toxin inhibits release of neurotransmitter, causing muscle paralysis. There are several serotypes, A to G, all of molecular weight about 150,000. The protein exists as a single chain or or as two chains, with two disulfide linkages. In a recent investigation on intracellular action of neurotoxins it was reported that type B neurotoxin can inhibit the release of Ca++-activated [3H] norepinephrine only if the disulfide bonds are reduced. In order to investigate possible structural changes in the toxin upon reduction of the disulfide bonds, we have prepared two-dimensional crystals of reduced type B neurotoxin. These two-dimensional crystals will be compared with those of the native (unreduced) type B toxin.

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Serum IGA subclass concentrations and functional antibody activity in intensive care patients (ICU)

Immunology Letters ◽

10.1016/s0165-2478(97)87398-1 ◽

1997 ◽

Vol 56 (1-3) ◽

pp. 140

Author(s):

J Miles

Keyword(s):

Intensive Care ◽

Antibody Activity ◽

Intensive Care Patients ◽

Serum Iga

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Specific Cross-reaction of IgG Anti-phospholipid Antibody with Platelet Glycoprotein IIIa

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650238 ◽

1996 ◽

Vol 75 (01) ◽

pp. 168-174 ◽

Cited By ~ 13

Author(s):

Shigeru Tokita ◽

Morio Arai ◽

Naomasa Yamamoto ◽

Yasuhiro Katagiri ◽

Kenjiro Tanoue ◽

...

Keyword(s):

Platelet Aggregation ◽

Heparan Sulfate ◽

Disulfide Bonds ◽

Dextran Sulfate ◽

Human Platelets ◽

Platelet Glycoprotein ◽

Activated Platelets ◽

Fibrinogen Binding ◽

Dose Dependent Fashion ◽

Glycoprotein Iiia

SummaryTo study the pathological functions of anti-phospholipid (anti-PL) antibodies, we have analyzed their effect on platelet function. We identified an IgG anti-PL mAb, designated PSG3, which cross-reacted specifically with glycoprotein (GP) IIIa in human platelets and inhibited platelet aggregation. PSG3 bound also to certain polyanionic substances, such as double-stranded DNA, heparan sulfate, dextran sulfate and acetylated-LDL, but not to other polyanionic substances. The binding of PSG3 to GPIIIa was completely inhibited by heparan sulfate and dextran sulfate, indicating that PSG3 recognizes a particular array of negative charges expressed on both GPIIIa and the specified polyanionic substances. Since neither neuraminidase- nor endoglycopeptidase F-treatment of GPIIIa had any significant effect on the binding of PSG3, this array must be located within the amino acid sequence of GPIIIa but not in the carbohydrate moiety. Reduction of the disulfide bonds in GPIIIa greatly reduced its reactivity, suggesting that the negative charges in the epitope are arranged in a particular conformation. PSG3 inhibited platelet aggregation induced by either ADP or collagen, it also inhibited fibrinogen binding to activated platelets in a dose-dependent fashion. PSG3, however, did not inhibit the binding of GRGDSP peptide to activated platelets. These results suggest that the PSG3 epitope on GPIIIa contains a particular array of negative charges, and possibly affects the fibrinogen binding to GPIIb/IIIa complex necessary for platelet aggregation.

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