Limits...
cDNA sequence and Fab crystal structure of HL4E10, a hamster IgG lambda light chain antibody stimulatory for γδ T cells.

Verdino P, Witherden DA, Podshivalova K, Rieder SE, Havran WL, Wilson IA - PLoS ONE (2011)

Bottom Line: We generated antibody HL4E10 as a probe to identify novel costimulatory molecules on the surface of γδ T cells which lack the traditional αβ T cell co-receptors CD4, CD8, and the costimulatory molecule CD28.The crystal structure of the HL4E10 Fab at 2.95 Å resolution reveals a rigid combining site with pockets faceted by solvent-exposed tyrosine residues, which are structurally optimized for JAML binding.As the HL4E10 antibody is uniquely costimulatory for γδ T cells, humanized versions thereof may be of clinical relevance in treating γδ T cell dysfunction-associated diseases, such as chronic non-healing wounds and cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, The Scripps Research Institute, La Jolla, California, United States of America.

ABSTRACT
Hamsters are widely used to generate monoclonal antibodies against mouse, rat, and human antigens, but sequence and structural information for hamster immunoglobulins is sparse. To our knowledge, only three hamster IgG sequences have been published, all of which use kappa light chains, and no three-dimensional structure of a hamster antibody has been reported. We generated antibody HL4E10 as a probe to identify novel costimulatory molecules on the surface of γδ T cells which lack the traditional αβ T cell co-receptors CD4, CD8, and the costimulatory molecule CD28. HL4E10 binding to γδ T cell, surface-expressed, Junctional Adhesion Molecule-Like (JAML) protein leads to potent costimulation via activation of MAP kinase pathways and cytokine production, resulting in cell proliferation. The cDNA sequence of HL4E10 is the first example of a hamster lambda light chain and only the second known complete hamster heavy chain sequence. The crystal structure of the HL4E10 Fab at 2.95 Å resolution reveals a rigid combining site with pockets faceted by solvent-exposed tyrosine residues, which are structurally optimized for JAML binding. The characterization of HL4E10 thus comprises a valuable addition to the spartan database of hamster immunoglobulin genes and structures. As the HL4E10 antibody is uniquely costimulatory for γδ T cells, humanized versions thereof may be of clinical relevance in treating γδ T cell dysfunction-associated diseases, such as chronic non-healing wounds and cancer.

Show MeSH

Related in: MedlinePlus

Crystal structure of the HL4E10 Fab.(A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3101210&req=5

pone-0019828-g004: Crystal structure of the HL4E10 Fab.(A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.

Mentions: (A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.


cDNA sequence and Fab crystal structure of HL4E10, a hamster IgG lambda light chain antibody stimulatory for γδ T cells.

Verdino P, Witherden DA, Podshivalova K, Rieder SE, Havran WL, Wilson IA - PLoS ONE (2011)

Crystal structure of the HL4E10 Fab.(A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3101210&req=5

pone-0019828-g004: Crystal structure of the HL4E10 Fab.(A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.
Mentions: (A) Cartoon representation of the superimposition of the two HL4E10 Fab structures in the asymmetric unit. The two HL4E10 Fabs (LH and AB) are shown in dark and light gray, respectively. The CDR loops are color coded as in Fig. 1&2: CDR L1 yellow, CDR L2 cyan, CDR L3 orange, CDR H1 blue, CDR H2 pink, CDR H3 green. The Cα atoms of Fab LH and Fab AB superimpose with an r.m.s.d. of 0.63 Å. (B) Superimposition of the combining sites of HL4E10 Fab LH (CDR loops colored) and Fab AB (CDR loops gray) (in a similar orientation to Fig. 4C) reveals a rigid assembly without significant conformational differences. (C) Wall-eyed stereo representation of the molecular interactions which rigidify the HL4E10 CDR loops and lock the side chains in conformations predefined for high affinity ligand binding. For example, hydrogen bonds (black), CH-π interactions (grey), and hydrophobic stacking interactions occur at the interface of CDR L3 with CDRs H3, H2, and H1.

Bottom Line: We generated antibody HL4E10 as a probe to identify novel costimulatory molecules on the surface of γδ T cells which lack the traditional αβ T cell co-receptors CD4, CD8, and the costimulatory molecule CD28.The crystal structure of the HL4E10 Fab at 2.95 Å resolution reveals a rigid combining site with pockets faceted by solvent-exposed tyrosine residues, which are structurally optimized for JAML binding.As the HL4E10 antibody is uniquely costimulatory for γδ T cells, humanized versions thereof may be of clinical relevance in treating γδ T cell dysfunction-associated diseases, such as chronic non-healing wounds and cancer.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, The Scripps Research Institute, La Jolla, California, United States of America.

ABSTRACT
Hamsters are widely used to generate monoclonal antibodies against mouse, rat, and human antigens, but sequence and structural information for hamster immunoglobulins is sparse. To our knowledge, only three hamster IgG sequences have been published, all of which use kappa light chains, and no three-dimensional structure of a hamster antibody has been reported. We generated antibody HL4E10 as a probe to identify novel costimulatory molecules on the surface of γδ T cells which lack the traditional αβ T cell co-receptors CD4, CD8, and the costimulatory molecule CD28. HL4E10 binding to γδ T cell, surface-expressed, Junctional Adhesion Molecule-Like (JAML) protein leads to potent costimulation via activation of MAP kinase pathways and cytokine production, resulting in cell proliferation. The cDNA sequence of HL4E10 is the first example of a hamster lambda light chain and only the second known complete hamster heavy chain sequence. The crystal structure of the HL4E10 Fab at 2.95 Å resolution reveals a rigid combining site with pockets faceted by solvent-exposed tyrosine residues, which are structurally optimized for JAML binding. The characterization of HL4E10 thus comprises a valuable addition to the spartan database of hamster immunoglobulin genes and structures. As the HL4E10 antibody is uniquely costimulatory for γδ T cells, humanized versions thereof may be of clinical relevance in treating γδ T cell dysfunction-associated diseases, such as chronic non-healing wounds and cancer.

Show MeSH
Related in: MedlinePlus