Structural aspects of human IgM antibodies expressed in chronic B lymphocytic leukemia.
BACKGROUND
Malignant B cells from patients with chronic B lymphocytic leukemia (B CLL) generally express both surface IgM and the pan T cell antigen CD5, a characteristic of the B1 population of B lymphocytes. The IgM on the surface of these B CLL cells is frequently polyreactive with respect to its capacity to recognize multiple structurally dissimilar antigens (Ag).


OBJECTIVE
To understand the structural characteristics of the polyreactive binding sites of human IgM molecules expressed on B CLL cells by: (1) analyzing the nucleotide and protein sequences of the variable (V) domains of five IgM molecules expressed in cases of B CLL and; (2) utilizing these sequences to generate three-dimensional (3D) models of Fv (VL - VH) molecules.


METHODS
Peripheral blood leukocytes obtained from five cases of B CLL were tested for polyreactive binding properties by assessing their capacity to bind mouse IgG by indirect immunofluorescence. The V region genes of light and heavy chains were amplified using the polymerase chain reaction, subsequently cloned and their nucleotide sequences obtained. Translated amino acid sequences of the V domains were used to generate homology models of the Fv molecules.


RESULTS
Low affinity binding of mouse IgG was demonstrated for all B CLL samples examined, confirming the polyreactive nature of the IgM expressed on these cells. There was an absence or minimal mutation within V region genes when compared to germline Ig genes. Junctional diversity was not observed for VL regions, although truncations and insertions were frequent in D minigenes of VH regions. The binding sites were predicted to form either relatively flat surfaces with occasional protrusions or cavities at the VL - VH domain interface. Aromatic side chains covered a large proportion of the potential binding surfaces in the models of B CLL Fv components.


CONCLUSIONS
Primary DNA sequences can be categorized as germline, suggesting that the B cells involved in B CLL are germline or naive in origin. The medium to large HCDR3s provide the majority of probable contact residues for antigens. While prominent aromatic residues are likely to engage in binding patterns which are conserved (e.g. mouse Ig reactivity), the diverse binding sites predicted for B CLL-derived IgMs also have properties which are conducive to polyreactive antigen binding.
Ramsland
P A
PA
Immunobiology Unit, University of Technology, Sydney, NSW, Australia.
Brock
C R
CR
Moses
J
J
Robinson
B G
BG
Edmundson
A B
AB
Raison
R L
RL
D000595:Amino Acid Sequence; D000818:Animals; D000912:Antibodies, Neoplasm / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D000915:Antibody Affinity; D001483:Base Sequence; D001666:Binding Sites, Antibody; D018956:CD5 Antigens / Q000276:immunology; D005803:Genes, Immunoglobulin; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000276:immunology; D007147:Immunoglobulin Light Chains / Q000737:chemistry / Q000276:immunology; D007075:Immunoglobulin M / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D015451:Leukemia, Lymphocytic, Chronic, B-Cell / Q000276:immunology*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation
D000912:Antibodies, Neoplasm; D018956:CD5 Antigens; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region