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* Department of Microbiology, Immunology and Molecular Genetics and Molecular Biology Institute, University of California, Los Angeles, CA 90095; and
Department of Pediatrics, Hokkaido University School of Medicine, Sapporo, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain and
the L chain when Cys220 is followed by Arg221,
but not when Cys220 is followed by Pro221,
suggesting that the Cys in CH1 might be involved in
disulfide bonding to the L chain. However, here we show that covalent
assembly of the H and L chains in IgA2(n) requires hinge-proximal
Cys241 and Cys242 in CH2 and not
Cys196 or Cys220 in CH1. Using
pulse-chase experiments, we have demonstrated that wild-type IgA2(n)
with Arg221 and Cys241 and Cys242
assembles through a disulfide-bonded HL intermediate. In contrast, the
major intermediate for IgA2 m(1) with Pro221 assembly was
H2 even though both Cys241 and
Cys242 were present. Only a small fraction of IgA2 m(1)
assembles through disulfide-bonded HL. Overall, our studies indicate
that for IgA2 covalent assembly of the H and L chains requires the
hinge-proximal cysteines in CH2 and that the structure of
CH1 influences the efficiency with which this covalent bond
forms. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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There are two subclasses of human IgA, IgA1 and IgA2, with IgA2 existing as several allotypes, IgA2m(1), IgA2m(2), and IgA2(n) (5). The different forms of human IgA differ in their H and L chain disulfide-bonding pattern. In IgA1, IgA2m(2), and IgA2(n), a disulfide bond links the L chain to the H chain. In contrast, the major form of IgA2m(1) lacks disulfide bonds between the H and L chain, although recent studies have shown that a small amount of HL disulfide-linked IgA2m(1) is made (2).
Structural analysis has shown that for IgA1,
Cys145 and Cys204 in
CH1, Cys266 and
Cys323 in CH2, and
Cys369 and Cys432 in
CH3 form the intradomain disulfide bonds
characteristic of the Ig fold (Fig. 1
)
(6, 7, 8). In IgA1, Cys196 and
Cys200 in CH1 and
Cys242 and Cys299 in
CH2 form additional intradomain disulfide bonds
with the remaining Cys residues forming covalent bonds with other
chains: 133 to the L chain, 241 and 301 to the H chain, 311 to the
secretory component, and 471 to the J chain (9). A similar
structure has been proposed for IgA2m(1) except there is no disulfide
bond with the L chain since Cys133 is now Asp
(Fig. 1) (10). Although IgA2m(2) and IgA2(n) also
lack a Cys at 133, they form covalent bonds with the L chain. An
unresolved question is which Cys residues within IgA2m(2) and IgA2(n)
form the bond with the L chain.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Restriction endonucleases and molecular cloning enzymes were obtained from either New England Biolabs (Beverly, MA) or Stratagene (La Jolla, CA). [35S]Methionine was obtained from ICN Research Products (Costa Mesa, CA). The Sp2/0 myeloma cells were cultured in IMDM containing 5% bovine calf serum (BCS)3 (HyClone Laboratories, Logan, UT).
Engineering of 2m(1) and mutants of IgA2(n) by site-directed
mutagenesis using PCR
The 2(n) gene (5) in
Bluescript II (Stratagene, La Jolla, CA) was used as a template to
generate 2m(1) by mutating Ser212
and Arg221 in 2(n) to
Pro212 and Pro221,
respectively. Two PCR products spanning CH1 were
generated: The first product was generated using primer 1 and primer 2
containing the two mutations shown below. The second mutation generated
an EcoRI site. The second product was generated using primer
3, the reverse complement of primer 2, and primer 4 that primes in the
intron between the CH1 and
CH2 3' to a SacII site. The PCR
products were sequenced and the SalI-SacII
fragment containing the CH1 of IgA2m(1) was
cloned into the 2(n) gene to yield the
2m(1) gene.
To generate the cysteine mutants in CH1, primers 1 and 5 containing the EcoRI site were used to generate a PCR product including C196S. Primers 6 and 4 were used to generate the PCR product with the C220S mutation. To generate the C199, 220S double mutant, a SalI-EcoRI fragment containing the C196S mutation was ligated to the EcoRI-SacII fragment containing the C220S mutation. To generate the hinge cysteine mutants, primers 1 and 7 and primers 1 and 8 containing either of the C241S or C242S mutations were used. The PCR products were cloned into the TA vector, sequenced, and cloned into BlueScript+ containing the wild-type IgA2(n). The complete constant regions were cloned into the pSV2gpt vectors containing the dansyl variable regions (11, 12). The constant regions in the expression vectors were sequenced again to confirm the presence of the mutations. Primers used were: 1, 5'-AACAGCTATGACCATG-3'; 2, 5'-CCTCTGACCTGGGCAGGGCACAGTCACATCCTGGCTGGGAATTCGTGTAG-3'; 3, reverse complement of primer 2; 4, 5'-CTGTCCACCACCACCTCCTGGGG-3'; 5, 5'-GCTGGAATTCGTGTAGTGCTTCACGTGGCATGTCACGGACTTGCCGTCTGGGGACTGTGTGGCCGG-3'; 6, 5'-TACACGAATTCCAGCCAGGATGTGACTGTGCCCTCCCGAGGTCAGAGGG-3'; 7, 5'-TCCTCGAGGGCCGGGCGGTGCAGCGACAGTCGGGGGTGGGAGCATGGGGGAGGTGGGGGAAC-3'; and 8, 5'-TCCTCGAGGGCCGGGCGGTGCAGCGACAGTCGGGGGTGGCAGGATGGGGGAGGTGGGGGAAC-3'.
Expression of the wild-type and mutant chimeric H chains in Sp2/0 myeloma cells
The H chain expression vector was transfected into Sp2/0 cells
expressing the dansyl-specific chimeric 
L chain gene by
electroporation (12). Approximately 6 x
106 cells were washed in cold 0.02 M PBS (pH 6.8)
and incubated on ice for 10 min with 15 µg of DNA. Cells were pulsed
with an electric field of 200 V and 960 µF in a Gene Pulser apparatus
(Bio-Rad, Richmond, CA), washed once, resuspended in 12 ml of IMDM
containing 10% FCS, 100 µg/ml gentamicin (Life Technologies, Grand
Island, NY), and 100 U/ml nystatin (Life Technologies) and plated in
96-well tissue culture plates at 125 µl/well. After 2 days of growth,
an equal volume of medium containing 15 µg/ml hypoxanthine, 250
µg/ml xanthine, and 6 µg/ml mycophenolic acid was added to the
wells to select for mycophenolic acid-resistant colonies. After 2 wk,
the surviving colonies were screened for Ab production by ELISA
using microtiter plates coated with dansyl coupled to BSA with bound Ab
detected by alkaline phosphatase-conjugated goat antiserum to human
L chain (Sigma-Aldrich, St. Louis, MO). Color was developed by adding
p-nitrophenyl phosphate (Sigma-Aldrich), and the absorbance
at 410 nm was determined in a microplate reader (MR 700; Dynatech
Laboratories, Chantilly, VA). Clones producing the highest quantities
of Ab were expanded in IMDM containing 10% (v/v) BCS.
Large-scale cell culture and purification of IgA Abs
Transfectomas were cultured in roller bottles (BD Labware, Lincoln Park, NJ) in IMDM supplemented with 1% BCS (HyClone Laboratories) and 6 mM L-glutamine (Life Technologies). Supernatants were filtered to remove any cells and cell debris and supplemented with 10 mM phosphate buffer (pH 6.8), 0.45 NaCl, 0.02 M EDTA, and 0.02% NaN3. Proteins were purified by affinity chromatography using AH-Sepharose beads coupled with the dansyl isomer 2-dimethylaminonapthalene-5-sulfonyl chloride (Molecular Probes, Eugene, OR). Bound Abs were eluted with N-(5-carboxy-pentyl)-2-dimethyl-aminonaphthyl-5-sulfonamide and concentrated and the hapten was removed by extensive dialysis against Tris buffer (pH 7.8) containing 0.45 M NaCl and 0.02% NaN3. The concentration of proteins was determined using the bicinchoninic acid assay (Pierce, Rockford, IL) and was confirmed by intensity comparison with an Ig standard of known concentration following SDS-PAGE and staining with Coomassie blue.
Biosynthetic labeling and pulse-chase analysis
To biosynthetically label transfectants with [35S]methionine, 2 x 106 cells were washed twice, incubated at 37°C for 30 min in methionine-free medium (Mediatech, Washington, DC), and then incubated for 16 h with 1 ml of methionine-free medium containing 1% (v/v) FCS and 12.5 µCi of [35S]methionine. For pulse-chase experiments, 2 x 107 cells were washed twice, incubated in 0.5 ml of methionine-free medium at 37° for 30 min, and then pulsed by adding 125 µCi of [35S]methionine and incubating at 37°C. After a 5-min incubation, the chase was initiated by adding 10 ml of medium containing 10% BCS and 3 mg/ml (100x) of unlabeled methionine. At various times after the initiation of the chase, 1-ml aliquots of cells were removed and cooled on ice. Cells were pelleted by centrifuging at 13,000 x g for 30 s in the cold and lysed by incubation on ice for 30 min in 0.5 ml of 10 mM Tris buffer (pH 7.4) containing 1% (v/v) Nonidet P-40, 0.4% (v/v) deoxycholate, and 66 mM EDTA. The lysates were centrifuged at 13,000 x g for 5 min in the cold to remove any unlysed cells and cell debris.
To immunoprecipitate cytoplasmic IgA, lysates were incubated for 1
h on ice with 5 µl of rabbit anti-human -chain (Sigma) and 2.5
µl of rabbit anti-human Fab followed by incubation for 10 min on
ice with 75 µl of 10% fixed Staphylococcus aureus cells
with surface protein A (IgG Sorb; Enzyme Center, Malden, MA). The
bacteria with bound immune complexes were pelleted at 13,000 x
g for 1 min through a 1-ml layer of 30% sucrose in 10 mM
Tris buffer (pH 7.4) containing 0.5% Nonidet P-40, 0.2% deoxycholate,
33 mM EDTA, and 0.15% SDS. The pellet was washed twice with 10 mM Tris
buffer (pH 7.4) containing 1% Nonidet P-40, 0.4% deoxycholate, 66 mM
EDTA, and 0.3% SDS and once with H2O. The
pellets were resuspended in electrophoresis sample buffer, placed in a
boiling water bath for 2 min, and centrifuged at 13,000 x
g for 2 min and the supernatants were analyzed by
SDS-PAGE.
Two-dimensional gel analysis
The pattern of disulfide bonding in IgA was determined by two-dimensional "diagonal" electrophoresis (13). The proteins were separated by SDS-PAGE in the first dimension in 5% phosphate gels under nonreducing conditions. To determine the composition of the bands observed in 5% phosphate gels, the lanes were excised and reduced in 0.32 M DTT in sample buffer for 60 min at room temperature. The lanes containing the reduced proteins were placed above separating 12.5% Tris-glycine gels containing SDS and sealed with 1% (w/v) agarose in sample buffer. The reduced proteins were then separated by electrophoresis in the second dimension. The gels were fixed, dried, and exposed to Kodak XAR-5 film.
Western blotting
Five micrograms of wild-type or mutant IgA2(n) were separated by
SDS-PAGE in phosphate-buffered 5% gels and transferred to a Millipore
Immobilon-P membrane (Millipore, Bedford, MA) according to the method
of Towbin et al. (14). Nonspecific sites were blocked by
incubating the membrane for 2 h at room temperature in PBS
containing 3% (w/v) BSA. The -chain was detected by incubating for
1 h at room temperature with goat anti-human -chain
(Zymed, San Francisco, CA) diluted 1/5000 (v/v) in PBS containing 1%
BSA. The bound goat Abs were detected following incubation for 1 h
at room temperature with rabbit anti-goat (Sigma-Aldrich)
conjugated to HRP diluted 1/5000 (v/v) in PBS containing 1% BSA,
washing, and then exposure to ECL reagent (Amersham, Buckingham, U.K.).
Nitrocellulose was exposed to Kodak Hyperfilm MP for 15 s. The L
and J chains were detected using goat anti-human -chain
conjugated to HRP (diluted 1/10, 000) and rabbit anti-human J chain
kindly provided by Dr. K. Kobayashi (Hokkaido University School of
Medicine, Sapporo, Japan; diluted 1:10, 000), followed by donkey
anti-rabbit conjugated to HRP (Jackson ImmunoResearch Laboratories,
West Grove, PA), respectively. For sequential blots, bound Abs were
removed by incubating the blots for 10 min at 60°C in 60.4 mM
Tris-HCl (pH 6.7) containing 100 mM 2-ME and 2% SDS. The efficiency of
removal of the Abs was confirmed by exposure to film.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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IgA2 exists with three alternate H chains: 2m(1), 2m(2), and
2(n) (5). The CH1 domains of
IgA2(n) and IgA2m(2) are identical and differ from 2m(1) at two
positions: they have Ser212 and
Arg221 whereas 2m(1) has
Pro212 and Pro221 (Fig. 1
).
The CH1 domain of IgA2 has only four cysteine
residues and lacks Cys133 that forms the
disulfide bond with the L chain in IgA1. Nevertheless, only IgA2m(1)
fails to efficiently form HL disulfides; in both IgA2m(2) and IgA2(n),
disulfides link H and L chains. Our earlier studies had indicated that
when Pro221 was changed to Arg in
CH1 in IgA2m(1), an HL disulfide formed,
suggesting that, like what is seen for IgA1, the HL disulfide for IgA2
was located within CH1 (2).
To further investigate the cysteine residues important for determining
the covalent attachment of the L chain in IgA2, mutants of 2(n) that
lacked Cys196, Cys220 (data
not shown), or both were generated. Surprisingly, the removal of these
Cys residues did not impact the formation of HL disulfide-bonded
molecules, and in the absence of both Cys196 and
Cys220 covalently assembled IgA2(n) was seen
(Fig. 2) with most of the protein present
in the secretions as an
H4L4J dimer with some
H2L2 and free L chain also
observed. The mutant protein did not differ significantly from
wild-type IgA2(n) in its assembly pattern, indicating that neither
Cys196 nor Cys220 are
required for disulfide bonding with L chains.
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). Multiple molecular forms were isolated from the secretions of the
two mutants. Especially noteworthy in IgA2(n)C241S and IgA2(n)C242S was
the presence of multiple molecular species with covalently
attached J chain. A distinguishing feature of the two proteins was that
the free L chains in IgA2(n)C242S were mostly monomers, whereas
virtually all of the noncovalently attached L chain present in the
IgA2(n)C241S secretions was dimer. Even though most of the L chain is
noncovalently associated with the two mutants, there are minor species
of several sizes of assembled proteins with covalently attached L
chains.
To further examine their assembly state, IgA2(n)C241S and IgA2(n)C242S
were analyzed by two-dimensional gel electrophoresis, doing Western
blot analysis with anti-J chain and anti-L chain in the second
dimension (Fig. 3). At least five
different molecular species with covalently attached J chain are seen.
Especially striking is the covalent association of the J chain with
nonpolymeric forms, including H2. This is in
contrast to what is seen with IgA2(n) and IgA2(n)C196,220S in which the
J chain is seen covalently associated only with polymer (Fig. 2). A
small portion of the L chain is covalently attached to H chains; some
of these species also appear to have covalently associated J chain but
it is difficult to precisely define the composition of these polymers
since J and L chains are similar in molecular mass.
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Pulse-chase analysis of IgA2(n) confirmed that there was efficient
intracellular formation of an HL disulfide with nearly 45% of the
total material synthesized present as HL by 15 min (Fig. 4). Thus, the major pathway of assembly
was H + L 
HL H2L2
with a small amount of H2L seen. The
H2L2 molecules formed
polymers with associated J chain. The composition of the bands was
confirmed by separating the immunoprecipitate from the 120-min time
point under nonreducing conditions in the first dimension, reducing it
in situ and analyzing by SDS-PAGE in 12.5% gels in the second
dimension (Fig. 4B). Secretion commenced at 30 min and by
180 min, 7% and 10 of the total counts were secreted as dimeric
IgA and mIgA, respectively (data not shown).
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). Five minutes after
the chase, a band migrating at the position of HL was observed (Fig. 5A); this band increased to 18% of the total by 15 min and
then rapidly decreased. This suggested that in the presence of
Pro221 some HL disulfides form but that these
intermediates do not efficiently form
H2L2 molecules. By 30 min,
a band of Mr 130,000 constituted the major
intracellular intermediate and remained as the major intermediate until
180 min. A band of Mr 150,000,
comprising 18% of the total counts at 30 min, decreased in quantity
when secretion commenced at 180 min. When the immunoprecipitate from
120 min after the chase was analyzed by two-dimensional
electrophoresis, the band migrating at
Mr 130,000 in the first dimension
resolved into H chains, whereas the band at
Mr 150,000 resolved into major bands
corresponding to H and J chains (Fig. 5B). Therefore, the
band of Mr 130,000 consists of mostly
H2 and the band of Mr
150,000 consists predominately of H2J. Thus,
IgA2m(1) resembles IgA2(n)C241S and IgA2(n)C242S in that J chains are
covalently associated with H2 molecules. The
observation that there is an intracellular accumulation of
H2J in IgA2m(1) implicate it as a major
intermediate in the synthesis of dimeric IgA. Together these results
suggest that the major assembly pathway of IgA2m(1) is H +
HH2 with J chain association and further
polymerization. Compared with IgA2(n), IgA2 m(1) with
Pro211 is less efficient in the formation of
disulfide-linked HL molecules and only small amounts of covalently
associated L chain are seen in the polymers.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The H chain of human IgA has 14 (IgA2) or 15 (IgA1) cysteine residues. These cysteines form intradomain bonds increasing IgA’s stability and covalent bonds with other H chains, L chains, J chain, and secretory component. Within human IgG, cysteine residues within the hinge (IgG1) or CH1 (IgG2, 3, and 4) form the covalent bond with the L chain. Consistent with this bonding pattern, Cys133 within the CH1 domain of IgA1 has been found to be required for the covalent attachment of L chain and its mutation to the aspartic acid found at the same position in IgA2 results in molecules with only noncovalent interactions between the H and L chains (2). Molecular modeling predicts that Cys133 will fold into the proximity of the terminal cysteine of the L chain. These same studies showed that HL disulfide bond formation could be restored to IgA2m(1) by mutating the proline next to Cys220 to an arginine, suggesting that a cysteine within CH1 may be responsible for forming the HL disulfide in IgA2. However, we have now shown that neither Cys196 nor Cys220 in CH1 are required for the formation of the HL disulfide in IgA2; instead, absence of either Cys241 or Cys242 in CH2 disrupts the formation of this disulfide. Thus, either Cys241 or Cys242 form the HL disulfide bond and alterations in the conformation of CH1 by the presence of Pro220 affect the ability of this bond to form.
No crystal structure exists for IgA; however, molecular models of IgA1
have been published (15, 16). In these models,
displacement of the CH2 domains was required to
accommodate the glycosylation and disulfide-bonding pattern leading to
steric crowding around the N-terminal region of the hinge compared with
IgG1. The disulfide bonds also restrict the movement of the C-terminal
region of the hinge and N-terminal region of the
CH2. However, in IgA1 the extended proline-rich,
O-glycosylated hinge region allows the Fab regions greater
conformational freedom relative to the corresponding domains in IgG1
and places the L chain Cys used to disulfide bond with the H chain at a
distance from Cys241. In IgA2, the two Fabs are
separated from the CH2 by only nine amino acids.
Consequently, the two Fab regions would be closer together in
comparison to IgA1 (16) and the penultimate cysteine
residues in the two L chains may be positioned closer to each other
and/or in close proximity to the hinge cysteines. In IgA2m(1), the
presence of Pro221 could form a kink in the
-chains, thereby making disulfide bond formation of the L chain with
Cys241/242 more difficult.
It is interesting that loss of either Cys241 or Cys 242 disrupts HL disulfide bond formation, making it difficult to directly assign which Cys forms the bond. However, the disulfide-bonding patterns of IgA1 indicates that Cys242 and Cys299 form an intradomain bond and would therefore be in close proximity in the folded structure. Since Cys241 is next to Cys242, it would also be in close proximity to Cys299 in the folded Ab. Intramolecular disulfide bond formation between cysteine residues in close proximity would be expected to occur more easily than would the intermolecular disulfide between H and L which requires that two chains that are independently synthesized must noncovalently associate to bring the two Cys residues into close proximity for the formation of the disulfide. It therefore seems likely that if only one Cys is present at either 241 or 242 it would participate in the intradomain disulfide bond. Cys301 used to form the H-H disulfide, is oriented into the space between the CH2 domains (15), allowing a disulfide to form even when the Fab is in close proximity to CH2. Cys311 is exposed on the surface consistent with its proposed role in forming a disulfide bond with the secretory component.
L chains contain five cysteine residues, four of which participate in intradomain disulfide bonds while the fifth can form a covalent bond with H chains or another L chain. For L chains to be secreted, the carboxyl-terminal Cys must either be covalently linked to a H or L chain or paired with a free cysteine (17). It is interesting that for IgA2(n)C242S, the majority of the secreted L chain is free, whereas for IgA2(n)C241S, most of the secreted L chain is dimeric. It is possible that the noncovalently associated L chains assume different conformations depending on whether the intradomain bond is between Cys241 and Cys299 or Cys242 and Cys299. Presumably when the intradomain bond is between Cys242 and Cys299, the penultimate cysteines in the L chains are brought into close proximity and efficiently form inter-L chain disulfide bonds.
J chain is associated with polymeric IgA and IgM (18).
Analysis of human colostral IgA showed that dimers contained 1 mol of J
chain for every two monomer units while the tetramers contained one J
for every four monomers units (19). J chain
contains eight Cys residues, six of which are involved in intrachain
disulfide bridges while two are disulfide linked to two penultimate
-chain cysteines in an IgA dimer with the other penultimate Cys of
the -chain forming an intra-H chain bond (8).
Substitution of either of the Cys residues of the J chain involved in
disulfide bond formation with the -chain yielded IgA monomers with
covalent J chain (20). The disulfide structure is not
consistent with previous models of the J chain structure (21, 22) but suggests that the J chain is positioned between two IgA
dimers linked tail-to-tail. C3 with the
associated tail piece but not C2 is required
for J chain incorporation into IgA (23). It is therefore
puzzling as to why novel forms with covalent J chain are seen in both
IgA2(n)C241S and IgA2(n)C242S (Figs. 2 and 3). These forms cannot be
solely the consequence of a failure to covalently attach a L chain
since they are not present in IgA2m(1) (our unpublished
observations).
Traditionally, IgA2m(1) has been described as being a protein lacking
covalent bonds between H and L (24, 25, 26) while IgA1,
IgA2m(2) (27), and novel IgA (5) have
covalent HL disulfides. Our studies have shown that this is not
strictly true. For IgA2m(1), some HL disulfide bonds do form, albeit
only inefficiently (2), suggesting that in IgA2m(1) the
cysteine residue involved in forming the HL disulfide is only partially
accessible and the majority of the molecules fail to form this bond.
For IgA2(n), some noncovalently associated L chains are also secreted
(Fig. 2). Since the Abs were purified on Ag columns, this L chain was
associated with the H chain during purification. It is interesting that
in IgA2m(1) a significant amount of the noncovalent L chains are dimers
(2), whereas in IgA2(n) most of the noncovalent L chains
are monomers (Fig. 2), again suggesting that the noncovalent L chains
assume different orientations in the two molecules.
In IgA2, while the shorter hinge may restrict the movements of the Fab regions to access Ags, it provides a functional advantage by being resistant to bacterial IgA1 proteases. This may explain why IgA2 is more abundant in the intestinal secretions where most of the bacteria reside. In addition, IgA2m(2) and IgA2(n) with covalently linked H and L chains may be more stable than IgA2m(1) in the milieu of the mucosal secretions with varying pH and salt concentrations.
In summary, we have determined the covalent assembly pathways in IgA and identified some of the cysteine residues and the structures in IgA that influence disulfide bond formation in IgA. The covalent assembly of IgA is expected to be essential for it to function efficiently in the hostile environment of the secretions of the respiratory, gastrointestinal, and genito-urinary tracts. Since disulfide bonds among H, L, and J chains are a major force in stabilizing the three-dimensional structure of IgA, the knowledge gained from these studies will enable us to better design and produce efficient immunotherapeutics based on IgA.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Koteswara R. Chintalacharuvu, Department of Microbiology, Immunology and Molecular Genetics, 611 Charles Young Drive South, Paul Boyer Hall 519, University of California, Los Angeles, CA 90095-1489. E-mail address: kotec{at}lifesci.ucla.edu
3 Abbreviation used in this paper: BCS, bovine calf serum.
Received for publication May 31, 2002. Accepted for publication August 27, 2002.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain constant region gene sequences: a novel recombinant 2 gene. J. Immunol. 152:5299.[Abstract]
receptor interactions. J. Biol. Chem. 273:2260.-chain CH2 domain. J. Immunol. 159:244.[Abstract]
A-globulins (-A2) which lacks the disulfide bonds linking heavy and light chains. J. Exp. Med. 128:1223.[Abstract]
A2 globulins. Proc. Natl. Acad. Sci. 65:557.2 heavy chain of a human IgA2 immunoglobulin of the A2m(2) allotype. Proc. Natl. Acad. Sci. USA 75:966.This article has been cited by other articles:
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  K. R. Chintalacharuvu, P. D. Chuang, A. Dragoman, C. Z. Fernandez, J. Qiu, A. G. Plaut, K. R. Trinh, F. A. Gala, and S. L. Morrison Cleavage of the Human Immunoglobulin A1 (IgA1) Hinge Region by IgA1 Proteases Requires Structures in the Fc region of IgA Infect. Immun., May 1, 2003; 71(5): 2563 - 2570. [Abstract] [Full Text] [PDF]
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