PDBsum entry 2ipk

Immune system

PDB id: 2ipk

Contents

Protein chains

179 a.a. *

190 a.a. *

14 a.a. *

232 a.a. *

Waters ×419

* Residue conservation analysis

PDB id:

2ipk

Name:

Immune system

Title:

Crystal structure of the mhc class ii molecule hla-dr1 in complex with the fluorogenic peptide, acpkxvkqntlklat (x=3-[5-(dimethylamino)-1,3- dioxo-1,3-dihydro-2h-isoindol-2-yl]-l-alanine) and the superantigen, sec3 variant 3b2

Structure:

Hla class ii histocompatibility antigen, dr alpha chain. Chain: a. Fragment: extracellular domain, residues 26-207. Synonym: mhc class ii antigen dra. Engineered: yes. Hla class ii histocompatibility antigen, drb1-1 beta chain. Chain: b. Fragment: extracellular domain, residues 30-219. Synonym: mhc class i antigen drb1 1, Dr-1, dr1.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: hla-dra. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008. Gene: hla-drb1. Synthetic: yes. Influenza a virus.

Resolution:

2.30Å R-factor: 0.203 R-free: 0.224

Authors:

T.T.Nguyen,L.J.Stern

Key ref:

P.Venkatraman et al. (2007). Fluorogenic probes for monitoring peptide binding to class II MHC proteins in living cells. Nat Chem Biol, 3, 222-228. PubMed id: 17351628 DOI: 10.1038/nchembio868

Date:

12-Oct-06 Release date: 13-Mar-07

Protein chain

P01903  (DRA_HUMAN) -  HLA class II histocompatibility antigen, DR alpha chain from Homo sapiens

Seq: 254 a.a.

Struc: 179 a.a.

Protein chain

P01911  (2B1F_HUMAN) -  HLA class II histocompatibility antigen, DRB1 beta chain from Homo sapiens

Seq: 266 a.a.

Struc: 190 a.a.*

Protein chain

A8CDU0  (A8CDU0_9INFA) -  Hemagglutinin (Fragment) from Influenza A virus

Seq: 104 a.a.

Struc: 13 a.a.*

Protein chain

P0A0L5  (ENTC3_STAAU) -  Enterotoxin type C-3 from Staphylococcus aureus

Seq: 266 a.a.

Struc: 232 a.a.*

* PDB and UniProt seqs differ at 18 residue positions (black crosses)

DOI no: 10.1038/nchembio868

Nat Chem Biol 3:222-228 (2007)

PubMed id: 17351628

Fluorogenic probes for monitoring peptide binding to class II MHC proteins in living cells.

P.Venkatraman, T.T.Nguyen, M.Sainlos, O.Bilsel, S.Chitta, B.Imperiali, L.J.Stern.

ABSTRACT

A crucial step in the immune response is the binding of antigenic peptides to major histocompatibility complex (MHC) proteins. Class II MHC proteins present their bound peptides to CD4(+) T cells, thereby helping to activate both the humoral and the cellular arms of the adaptive immune response. Peptide loading onto class II MHC proteins is regulated temporally, spatially and developmentally in antigen-presenting cells. To help visualize these processes, we have developed a series of novel fluorogenic probes that incorporate the environment-sensitive amino acid analogs 6-N,N-dimethylamino-2-3-naphthalimidoalanine and 4-N,N-dimethylaminophthalimidoalanine. Upon binding to class II MHC proteins these fluorophores show large changes in emission spectra, quantum yield and fluorescence lifetime. Peptides incorporating these fluorophores bind specifically to class II MHC proteins on antigen-presenting cells and can be used to follow peptide binding in vivo. Using these probes we have tracked a developmentally regulated cell-surface peptide-binding activity in primary human monocyte-derived dendritic cells.

Selected figure(s)

Figure 1.
(a) Chemical structures of DANA, 4-DAPA and 6-DMNA. (b) In the crystal structure of the DR1–HA peptide complex, a tyrosine residue is buried deep into the hydrophobic P1 pocket^5. DANA, 4-DAPA and 6-DMNA were modeled in silico into this pocket in place of tyrosine. Residues shown lining the pocket are (clockwise from upper right) Phe 54, Phe 32, Trp 43 (behind), Ile 7, Trp 153, Phe 48, Thr 90, Val 91, Tyr 83, Gly 86 and Val 85. The side chains of Asn 82 and His 81 (upper left) form hydrogen bonds with the peptide main chain at the mouth of the P1 pocket. Other residues lining the pocket but not shown are Phe 24, Phe 26 and Phe 48. (c) Binding of fluorogenic peptides to DR1 assessed using a competitive binding assay. DR1 was incubated with biotin–HA peptide and various concentrations of unlabeled inhibitor peptides, and biotin-HA binding was quantified by a sandwich ELISA assay using streptavidin alkaline phosphatase. Binding of Fmoc-(4-DAPA) was assessed to evaluate nonspecific binding of the fluorophore. IC[50] values for these and other peptides (Supplementary Table 1) were determined as described (Supplementary Methods).

Figure 3.
(a) (4-DAPA)-HA was incubated with HLA-DR1 as described for (4-DAPA)-RSMA[4]L, and the complex formed was purified by gel filtration. Fluorescence of DR-(4-DAPA)-HA was compared with that of the free peptide. (b) Activation of HA1.7 T-cell receptor hybridoma by antigen-presenting cells pulsed with HA (filled circles) or (4-DAPA)-HA (closed triangles) peptides. T-cell activation reported as counts per minute (c.p.m.) measured in a thymidine incorporation bioassay for IL-2 as secreted by activated T cells. Error bars indicate s.d. of triplicate measurements. (c–e) Crystal structure of (4-DAPA)-HA bound to DR1. (c) (4-DAPA)-HA peptide shown with surface representation of the DR1 peptide binding site, with the 4-DAPA side chain shown with yellow bonds extending down into the P1 pocket (top); unmodified HA peptide from the crystal structure of the DR1-HA-SEC (3B2) complex (PDB ID 1JWU) shown after alignment of MHC peptide binding domain, with tyrosine side chain at the P1 position shown with cyan bonds (bottom). (d) 2F[o] - F[c] omit map of the region around the P1 pocket with all residues shown removed from the model before map calculation. (e) Section through the P1 pocket, showing the HA peptide tyrosine side chain and the (4-DAPA)-HA fluorophore, along with the corresponding ordered water molecules, colored as in c. Panels c–e made using PyMOL^30.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Chem Biol (2007, 3, 222-228) copyright 2007.

Figures were selected by the author.