scholarly journals The sequence of addition of terminal sugars to an immunoglobulin A myeloma protein

1972 ◽  
Vol 126 (3) ◽  
pp. 751-754 ◽  
Author(s):  
N. J. Cowan ◽  
G B Robinson
Keyword(s):  
Immunoglobulin A ◽  
Myeloma Protein

Structure of immunoglobulin A. Cysteine-containing peptides of the α chain of an immunoglobulin A1 myeloma protein

Biochemistry ◽  
1973 ◽  
Vol 12 (25) ◽  
pp. 5186-5194 ◽  
Author(s):  
Enrique Mendez ◽  
Blas Frangione ◽  
Edward C. Franklin
Keyword(s):  
Immunoglobulin A ◽  
Myeloma Protein ◽  
Α Chain

scholarly journals Biosynthesis of immunoglobulin A (IgA). Secretion and addition of carbohydrate to monomer and polymer forms of a mouse myeloma protein

1973 ◽  
Vol 136 (3) ◽  
pp. 589-596 ◽  
Author(s):  
E. Della Corte ◽  
R. M. E. Parkhouse
Keyword(s):  
Plasma Cell ◽  
Molecular Species ◽  
Specific Antibody ◽  
Early Stage ◽  
Immunoglobulin A ◽  
Carbohydrate Composition ◽  
Myeloma Protein ◽  
J Chain ◽  
Co Precipitation ◽  
Mouse Myeloma

Cell suspensions of mouse plasma-cell tumour MOPC 315 secreting predominantly IgA (immunoglobulin A) monomer and dimer were incubated with radioactive leucine, mannose, galactose and fucose for various periods of time. The amounts of secreted and intracellular immunoglobulins were measured by co-precipitation with specific antibody, and the molecular species present were assessed by electrophoresis in polyacrylamide gels. Analysis of the secreted myeloma protein demonstrated that monomer and dimer IgA molecules are identical with respect to carbohydrate composition and rate of secretion. Within the cell, the myeloma protein is almost entirely accounted for by monomer units which either leave the cell as such or are polymerized with the addition of J chain close to the time of secretion. The results support the concept of a stepwise addition of carbohydrate residues to IgA immunoglobulin during the process of secretion. Similar patterns of carbohydrate assembly were found for the monomer or dimer molecules. Mannose residues are added at an early stage, whereas fucose is added close to the time of secretion. Galactose is also added early, but some may also be incorporated at a later stage. Control of IgA polymerization is considered unlikely to reflect regulation at the level of carbohydrate addition, and it is suggested that the critical controlling factor is the J chain.

scholarly journals The thiol groups of mouse immunoglobulin A. Incomplete formation of the Cα 1-domain disulphide bridge

1985 ◽  
Vol 225 (1) ◽  
pp. 113-125 ◽  
Author(s):  
S A Cockle ◽  
N M Young
Keyword(s):  
High Performance ◽  
Immunoglobulin A ◽  
Secretory Component ◽  
Reduced Form ◽  
Thiol Groups ◽  
Disulphide Bridge ◽  
Myeloma Protein ◽  
Sh Groups ◽  
Heavy Chains ◽  
Bond Model

The BALB/c IgA (immunoglobulin A) myeloma protein M167 contained on average 5.7 free SH groups per IgA dimer. These groups were preponderantly on the heavy chains and comprised two distinct populations: 3.3 exposed SH groups per dimer in the Fc region, and 2.4 buried SH groups per dimer in the Fd region, detectable o only after denaturation. To locate the cysteine residues involved, labelled peptides were purified from thermolysin digests of radioalkylated IgA by high-performance liquid chromatography. From the amino acid compositions of the peptides, the exposed thiol groups were assigned to Cys-307 in the C alpha 2 domain, which thus existed in the reduced form to an extent exceeding 80%. This residue may allow attachment of secretory component to dimer IgA in the mouse to proceed via thiol-disulphide exchange. The buried thiol groups were assigned to Cys-150 and Cys-208, in the C alpha 1 domain, each being in the reduced form to the extent of approx. 30%. This pair of residues would normally give rise to the characteristic intradomain disulphide bridge. It appears that disulphide formation is not a crucial event during folding of the C alpha 1 domain in IgA biosynthesis. The sequence in the region 140-151 was re-investigated, and residue 142 was shown to be serine, not cysteine, helping explain the lack of heavy-chain-light chain bonding in BALB/c mouse IgA. A disulphide-bond model for mouse IgA is proposed on the basis of these assignments and other features of the mouse alpha-chain sequence.

Subunit structure and number of combining sites of the immunoglobulin A myeloma protein produced by mouse plasmacytoma MOPC-315

Biochemistry ◽  
1971 ◽  
Vol 10 (24) ◽  
pp. 4359-4368 ◽  
Author(s):  
Brian J. Underdown ◽  
Ernest S. Simms ◽  
Herman N. Eisen
Keyword(s):  
Immunoglobulin A ◽  
Subunit Structure ◽  
Myeloma Protein

Immunoglobulin A. Arrangement of disulfide bridges in the hinge region of an immunoglobulin A1 human myeloma protein

Biochemistry ◽  
1973 ◽  
Vol 12 (25) ◽  
pp. 5195-5197 ◽  
Author(s):  
Carlota Wolfenstein-Todel ◽  
Frances Prelli ◽  
Blas Frangione ◽  
Edward C. Franklin
Keyword(s):  
Immunoglobulin A ◽  
Hinge Region ◽  
Disulfide Bridges ◽  
Myeloma Protein

scholarly journals Isolation of plasma membrane from protoplasts of Lolium multiflorum (ryegrass) endosperm cells

1982 ◽  
Vol 205 (3) ◽  
pp. 511-519 ◽  
Author(s):  
A Schibeci ◽  
G B Fincher ◽  
B A Stone ◽  
A B Wardrop
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Membrane Fraction ◽  
Plasma Membranes ◽  
Lolium Multiflorum ◽  
Immunoglobulin A ◽  
Specific Radioactivity ◽  
Centrifugal Forces ◽  
Myeloma Protein ◽  
Plasma Membrane Fraction

Plasma membranes have been isolated from protoplasts of suspension-cultured ryegrass (Lolium multiflorum) endosperm cells. The protoplast membrane is coated before cell disruption with murine myeloma protein J539, a galactose-binding immunoglobulin A. The plasma membrane is labelled with 125I by using chemically or enzymically catalysed iodination techniques, or, more conveniently, by using 125I-labelled myeloma protein J539, which enables the membrane to be simultaneously coated and labelled. Protoplast lysis is effected by gentle mechanical means after swelling in hypo-osmotic medium. The plasma-membrane fraction is recovered at low centrifugal forces by fractionation of cell lysates on a discontinuous sucrose/sorbitol gradient. The plasma-membrane fraction is enriched 96-fold on a protein basis with respect to the specific radioactivity of 125I-labeled myeloma protein J539 in the homogenate. Electron microscopy showed long membrane profiles often associated with one another.

Structure of immunoglobulin A. I. Interchain disulfide bridges of a γAl myeloma protein

Biochemistry ◽  
1971 ◽  
Vol 10 (22) ◽  
pp. 4140-4145 ◽  
Author(s):  
C. Wolfenstein ◽  
Blas Frangione ◽  
E. Mihaesco ◽  
E. C. Franklin
Keyword(s):  
Immunoglobulin A ◽  
Disulfide Bridges ◽  
Myeloma Protein

Small-angle neutron scattering studies of the conformation of myeloma protein MOPC315 and its Fab fragment, and the interaction with a monovalent dinitrophenyl hapten

1981 ◽  
Vol 211 (1185) ◽  
pp. 433-453 ◽  
Keyword(s):  
Immunoglobulin G ◽  
Small Angle ◽  
Mass Density ◽  
Immunoglobulin A ◽  
Molecular Volume ◽  
Fab Fragment ◽  
Cross Sectional ◽  
Myeloma Protein ◽  
Contrast Method ◽  
Angle Scattering

The first small-angle scattering study of an immunoglobulin A is reported. Neutron measurements have been made to determine conformational parameters of the mouse myeloma protein MOPC315 and to relate these to previous immunoglobulin G results. Use of the contrast method shows that the MOPC315 IgA molecule is not simply globular, that it has a dry volume of 220.0 ± 4.5 nm 3 corresponding to a mass density of 1.275 ± 0.025 g cm −3 and that its full and cross-sectional radii of gyration, corrected for con­centration dependence, are 7.97 ± 0.07 nm, 2.40 ± 0.08 nm and 1.33 ± 0.07 nm respectively. Similar study of its Fab fragment gives a dry molecular volume of 69.0 ± 0.7 nm 3 , a mass density of 1.285 ± 0.015 g cm −3 and un­corrected radii of gyration that are consistent with those of the parent and support an overall ‘ T ’ or ‘ Y ’ conformation in solution. Addition to satu­ration of a small monovalent dinitrophenyl hapten leaves the dry volume of the whole molecule unaltered, but may slightly lower one or more of its radii of gyration. The significance of this finding is discussed. Comparative studies with rabbit anti-dinitrophenyl immunoglobulin G antibody suggest a different initial conformation but similar consequences of hapten binding, which, if real, are probably unrelated to classical complement fixation.

scholarly journals The gross architecture of an antibody-combining site as determined by spin-label mapping

1977 ◽  
Vol 165 (2) ◽  
pp. 177-197 ◽  
Author(s):  
Brian J. Sutton ◽  
Peter Gettins ◽  
David Givol ◽  
Derek Marsh ◽  
Simon Wain-Hobson ◽  
...  
Keyword(s):  
Light Chain ◽  
Metal Binding ◽  
Spin Label ◽  
Immunoglobulin A ◽  
Variable Region ◽  
Spin Labels ◽  
Myeloma Protein ◽  
Polarity Profile ◽  
Resonance Relaxation ◽  
Hyperfine Splittings

1. A series of Dnp (dinitrophenyl) nitroxide spin labels was used to map the dimensions of the combining site of the Dnp-binding immunoglobulin A myeloma protein MOPC 315. The method compares the observed e.s.r. (electron-spin-resonance) hyperfine splittings with those calculated on the basis of different postulated motions for the spin label. The analysis is complicated by the sensitivity of the e.s.r. hyperfine splitting to the overall ‘tumbling’ time of the antibody–hapten complex and the polarity of the spin-label's environment. When these effects are considered quantitatively, it is then possible to determine the degree of mobility of each hapten which is allowed by the shape of the combining site. 2. The dinitrophenyl ring is rigidly held, and the depth of the site is 1.1–1.2nm and has lateral dimensions at the entrance to the site ≥0.6nm×0.9nm. The analysis of the results for spin-labelled haptens with chiral centres allows these lateral dimensions to be refined to 0.8nm and 1.1nm, and it is shown that the site is asymmetric with respect to the plane of the dinitrophenyl ring. 3. A polarity profile of the combining site was also obtained and a positively charged amino acid residue, possibly arginine-95L (light chain), was located at the entrance to the site. 4. The binding of Gd(III) to the antibody–hapten complexes results in quenching of the e.s.r. signal of the nitroxide. By using La(III) as a control, the paramagnetic contribution to the quenching is measured. 5. Analysis of the differential quenchings of the enantiomers of two five-membered nitroxide ring spin labels gives two possible locations of the metal-binding site. One of these is equidistant (0.7nm) from each of the three dinitrophenyl aromatic protons, and nuclear-magnetic-resonance relaxation studies, at 270MHz, on solutions of dinitrobenzene, Gd(III) and the Fv fragment (variable region of heavy and light chain) from protein MOPC 315 support this location for the metal site. 6. The e.s.r. and metal-binding data were then compared with the results of a model of the combining site constructed on the basis of framework invariance in immunoglobulins [Padlan, Davies, Pecht, Givol & Wright (1976) Cold Spring Harbor Symp. Quant. Biol.41, in the press]. The overall agreement is very good. Assignments of possible chelating groups for the metal can be made.

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