The Effects of Histidine Residue Modification on the Immune Precipitating Ability of Rabbit IgG

O'Brien, Roberta; Roeth, Philip; Thomson, Shannon; Bartell, G.; Easterbrook‐Smith, Simon B.

doi:10.1006/abbi.1994.1135

Cited by 3 publications

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“…By employing the specific histidine‐modifying reagent diethylpyrocarbonate (DEPC), 22 previous reports have established that histidine residues on the Fc portion of IgG play an important role in both, the binding to staphylococcal protein A and the homologous interactions among Fc regions required to the formation of insoluble IC. 23 , 24 To our knowledge no previous works have analysed the role of histidine residues on the Fc fragment of IgG in the binding of IC to FcγR. In an attempt to determine whether titration of these residues could account for the enhancing effect of acidic pH on IC binding to FcγR, we performed additional experiments using AIgG treated with DEPC.…”

Section: Discussionmentioning

confidence: 99%

Acidic pH increases the avidity of FcγR for immune complexes

et al. 1999

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SUMMARYThe interaction of immunoglobulin G (IgG) antibodies with FccR constitutes a critical mechanism through which IgG antibody effector functions are mediated. In the current work we have examined whether human neutrophil FccR exhibit pH dependence in their association with IgG. Binding assays were performed in culture medium adjusted to different pH values. It was found that the binding of either heat-aggregated human IgG (AIgG), soluble immune complexes (sIC) or IgGcoated erythrocytes (IgG-E) was markedly higher at pH 6 e 5 than at pH 7 e 3. This effect was not observed when saturation of FccR was achieved, suggesting that acidic pH increases the avidity of FccR for IC without modifying the total binding capacity. Similar results were observed for the binding of AIgG to either monocytes, natural killer (NK) or K562 cells, suggesting that acidic pH increases the avidity of both, FccRII and FccRIII. Additional experiments were performed to analyse whether the binding of IgG to FccRI also showed pH dependence. To this aim, we employed interferon-c-treated human neutrophils and mouse in¯ammatory macrophages, previously incubated with blocking antibodies directed to FccRII and FccRIII. Acidic pH did not enhance the binding of AIgG nor monomeric IgG under these experimental conditions. Further studies are required to determine whether the enhancement of FccR avidity for IC could be attributed to titration of histidine(s) residues on the Fc fragment of IgG.

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Section: Discussionmentioning

confidence: 99%

Acidic pH increases the avidity of FcγR for immune complexes

et al. 1999

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The formation of insoluble immune complexes between ovalbumin and anti-ovalbumin IgG occurs in at least two distinct phases dependent on reactant concentration and ionic strength

Gorgani

Easterbrook‐Smith

Altin

1996

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Diseas

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The mechanism regulating the formation of insoluble immune complexes (IIC) in serum in certain disease states is not well understood. Ovalbumin and rabbit anti-ovalbumin IgG was used to study the formation of IIC in vitro in a stirred reaction vessel; and the radii of IIC that formed was determined by light scattering techniques. Using an initial IgG concentration of 1 mg/ml at equivalence antigen:antibody ratio IIC formation was detected within 5 s, and the complexes increased in radii to approx. 100 nm after 20-30 s (phase 1). This was followed by a phase (phase 2) in which the complexes rapidly increased in radii to the point where Mie scattering was reached (approximately 200 nm). The time of onset of the second phase decreased with increasing initial IgG concentrations at a fixed antigen:antibody ratio; and was at a minimum at equivalence antigen:antibody ratio, but increased at both antigen and antibody excess ratios. Immune complexes formed using F(ab')2 fragment showed a similar pattern to those formed using IgG. A similar pattern was seen in the presence of the complement component C1q which potentiated IIC formation in phase 2, and human serum (1:10 dilution) which attenuated IIC formation in both phases. For complex formation using IgG and ovalbumin the presence of NaC1 at concentrations up to 0.6 M led to a progressive increase in the time of onset of phase 2; potencies of inhibition by other sodium halides followed the lyotropic series NaF < NaC1 < NaI. The results suggest that formation of IIC occurs in at least two distinct phases, and that the second phase leading to the generation of very large insoluble complexes is associated with a rapid polymerisation of the complexes by a mechanism that is not dependent on Fc:Fc interactions.

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Analysis of the Dynamics of Cryoaggregation by Light-Scattering Spectrometry

Stasio¹,

Bizzarri²,

Bove³

et al. 2003

Clinical Chemistry and Laboratory Medicine

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Cryoglobulins are proteins that precipitate at temperatures below 37 degrees C. Cold-induced precipitation of proteins may occur in vivo secondary to several important diseases, and lead to pathological manifestations involving different organs. Cryoprecipitation may be observed in vitro by exposing serum samples, supposed to contain cryoglobulins, to low temperatures, but this needs several days to occur. Protein-protein interactions leading to cryoprecipitation are still poorly understood and the knowledge of the underlying mechanism may be relevant to the understanding of the onset of pathological manifestations. Using light-scattering spectrometry, we studied cryoprecipitation occurring in vitro at different temperatures and cryoglobulin concentrations. We describe the kinetics of the cold-induced precipitation of mixed cryoglobulins, measured as increase in turbidity. The plots obtained demonstrate that the cryoprecipitation did not occur as a single-step reaction, but consisted of four distinct phases where both temperature and cryoglobulin concentration affected the immune complexes formation. Light scatter spectrometry may provide a simple, sensitive and rapid method for the detection of cryoglobulins.

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The Effects of Histidine Residue Modification on the Immune Precipitating Ability of Rabbit IgG

Cited by 3 publications

References 0 publications

Acidic pH increases the avidity of FcγR for immune complexes

Acidic pH increases the avidity of FcγR for immune complexes

The formation of insoluble immune complexes between ovalbumin and anti-ovalbumin IgG occurs in at least two distinct phases dependent on reactant concentration and ionic strength

Analysis of the Dynamics of Cryoaggregation by Light-Scattering Spectrometry

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