Characterizing the polymeric immunoglobulin binding region of human secretory component

Bakos, Mary Ann; Kurosky, A.; Goldblum, Randall M.

doi:10.1007/978-94-009-1848-1_84

Cited by 2 publications

(7 citation statements)

References 6 publications

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“…All the mSC-FLAG proteins mutated outside domain I bound with high affinity to IgA d , which corroborates with the previous finding that the first domain of rabbit and human pIgR and SC carries the most important information for ligand recognition (5,17,18). Nevertheless, although domain I of rabbit SC is both necessary and sufficient for binding IgA (17), domain III of bovine SC contributes to binding (21), and domains II-V of hSC add to its affinity for IgA (22). Using a competition assay, we found that the concentration for half-maximal inhibition (IC 50 ) was about 1 nM for wild type mSC and 1-6 nM for all mutants except mSC-FLAG:D1:FG (Table I) measured an IC 50 of 10 nM for the interaction taking place between either recombinant or milk-derived hSC and the same murine IgA d (16).…”

Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercasupporting

confidence: 88%

“…Thus, it appears that mSC has a higher affinity than hSC for murine IgA d . In the same setting an IC 50 of 3-30 nM (4,18,22) has been reported for the interaction of hSC isolated from milk with human IgA p . Furthermore, relative affinities for human IgA p of bovine and rabbit SC were in the range of 1 nM, whereas that of rat SC was about 40 nM (4).…”

Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercamentioning

confidence: 53%

“…In the same setting an IC 50 of 3-30 nM (4,18,22) has been reported for the interaction of hSC isolated from milk with human IgA p . Furthermore, relative affinities for human IgA p of bovine and rabbit SC were in the range of 1 nM, whereas that of rat SC was about 40 nM (4). Scatchard analysis yielded a K D of 10 nM for the binding of hSC to human IgA d (23), rabbit SC to rabbit IgA d (24), or rabbit IgA d to rabbit pIgR (25) and a K D of 2.5 nM for the binding of human IgA d to rabbit pIgR (26).…”

Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercamentioning

confidence: 53%

“…The complex is internalized at the basolateral surface of the epithelium and transcytosed to the apical plasma membrane, where the extracellular portion of the receptor called secretory component (SC) is released by proteolytic cleavage and remains bound to IgA within a complex termed secretory IgA (sIgA) (2,3). All three loops, referred to as B-C, D-E, and F-G in domain I are particularly implicated in binding to IgA (4,5); in addition, some residues of loop E-F in the opposite face appear to be important for IgA binding. In contrast, deletion of domains II and III or individual deletion of domain IV and V of rabbit pIgR did not prevent binding to IgA in a qualitative cell-ligand binding assay (5).…”

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confidence: 99%

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Mapping the Interaction Between Murine IgA and Murine Secretory Component Carrying Epitope Substitutions Reveals a Role of Domains II and III in Covalent Binding to IgA

Crottet¹,

Corthésy²

1999

Journal of Biological Chemistry

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We have identified sites for epitope insertion in the murine secretory component (SC) by replacing individual surface-exposed loops in domains I, II, and III with the FLAG sequence (Crottet, P., Peitsch, M. C., Servis, C., and Corthé sy, B. (1999) All three loops, referred to as B-C, D-E, and F-G in domain I are particularly implicated in binding to IgA (4, 5); in addition, some residues of loop E-F in the opposite face appear to be important for IgA binding. In contrast, deletion of domains II and III or individual deletion of domain IV and V of rabbit pIgR did not prevent binding to IgA in a qualitative cell-ligand binding assay (5). A natural variant of rabbit SC lacking domains II and III can bind to IgA, although only in a noncovalent fashion (6, 7). In human sIgA, a disulfide bridge within domain V is also involved in covalent binding to the C␣2 domain of IgA through a disulfide exchange mechanism (8 -10). Species variations in the level of covalency between pIgR and IgA have been reported (11,12). Thus, it is tempting to speculate that although not critical for the initial recognition of IgA, domains II to IV appear to position domain V such that disulfide exchange with IgA can take place.The mechanism for the selective recognition of IgA d over IgA m remains an additional open question. We have generated murine SC mutants with predicted exposed loops of domains I, II, and III replaced with the FLAG epitope (13) to evaluate their IgA binding properties. We show that although the affinity of these mutants for IgA and the selectivity for IgA d over IgA m are both preserved, covalent binding is lost in all mutants but one. Given the pinpointing strategy designed here, this suggests that domains II and III are essential to the proper positioning of domain V. However, because binding to the anti-FLAG mAb is preserved in both free and IgA d -bound reactive SC mutants, we postulate that minor structural changes occur upon IgA d -mSC interactions. Dimeric SC-FLAG mutants were shown to be as good IgA d binders as their monomeric counterparts. In the same assay, we found by direct comparison of mSC and pIgR mutants that mSC binds to IgA d with much reduced stringency. We conclude that mSC-FLAG mutants are useful 1) to study the topology of SC⅐IgA d complexes, 2) as short epitope carrier once combined with IgA d , 3) to tackle the binding specificity and stoichiometry of SC/pIgR IgA d complexes.

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Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercasupporting

confidence: 88%

Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercamentioning

confidence: 53%

Section: Biosynthesis Of Msc-flag Proteins In the Presence Of 2-mercamentioning

confidence: 53%

mentioning

confidence: 99%

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Mapping the Interaction Between Murine IgA and Murine Secretory Component Carrying Epitope Substitutions Reveals a Role of Domains II and III in Covalent Binding to IgA

Crottet¹,

Corthésy²

1999

Journal of Biological Chemistry

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“…However, the antigenized SC was very poorly secreted and mostly retained in the early secretion pathway. This study revealed a novel function for domain I in SC/pIgR secretion, besides its role in the initial, high affinity, binding to IgA d (11)(12)(13)(14)(15)(16)(17).…”

mentioning

confidence: 91%

Covalent Homodimers of Murine Secretory Component Induced by Epitope Substitution Unravel the Capacity of the Polymeric Ig Receptor to Dimerize Noncovalently in the Absence of IgA Ligand

Crottet¹,

Peitsch²,

Servis³

et al. 1999

Journal of Biological Chemistry

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Recombinant secretory immunoglobulin A containing a bacterial epitope in domain I of the secretory component (SC) moiety can serve as a mucosal delivery vehicle triggering both mucosal and systemic responses (Corth-é sy, B., Kaufmann, M., Phalipon, A., Peitsch, M., Neutra, M. R., and Kraehenbuhl, J.-P. (1996) J. Biol. Chem. 271, 33670 -33677). To load recombinant secretory IgA with multiple B and T epitopes and extend its biological functions, we selected, based on molecular modeling, five surface-exposed sites in domains II and III of murine SC. Loops predicted to be exposed at the surface of SC domains were replaced with the DYKDDDDK octapeptide (FLAG). Another two mutants were obtained with the FLAG inserted in between domains II and III or at the carboxyl terminus of SC. As shown by mass spectrometry, internal substitution of the FLAG into four of the mutants induced the formation of disulfide-linked homodimers. Three of the dimers and two of the monomers from SC mutants could be affinity-purified using an antibody to the FLAG, mapping them as candidates for insertion. FLAG-induced dimerization also occurred with the polymeric immunoglobulin receptor (pIgR) and might reflect the so-far nondemonstrated capacity of the receptor to oligomerize. By co-expressing in COS-7 cells and epithelial Caco-2 cells two pIgR constructs tagged at the carboxyl terminus with hexahistidine or FLAG, we provide the strongest evidence reported to date that the pIgR dimerizes noncovalently in the plasma membrane in the absence of polymeric IgA ligand. The implication of this finding is discussed in terms of IgA transport and specific antibody response at mucosal surfaces.

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Characterizing the polymeric immunoglobulin binding region of human secretory component

Cited by 2 publications

References 6 publications

Mapping the Interaction Between Murine IgA and Murine Secretory Component Carrying Epitope Substitutions Reveals a Role of Domains II and III in Covalent Binding to IgA

Mapping the Interaction Between Murine IgA and Murine Secretory Component Carrying Epitope Substitutions Reveals a Role of Domains II and III in Covalent Binding to IgA

Covalent Homodimers of Murine Secretory Component Induced by Epitope Substitution Unravel the Capacity of the Polymeric Ig Receptor to Dimerize Noncovalently in the Absence of IgA Ligand

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