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PDBsum entry 1df3
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Transport protein
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PDB id
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1df3
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Solution structure of a recombinant mouse major urinary protein.
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Authors
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C.Lücke,
L.Franzoni,
F.Abbate,
F.Löhr,
E.Ferrari,
R.T.Sorbi,
H.Rüterjans,
A.Spisni.
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Ref.
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Eur J Biochem, 1999,
266,
1210-1218.
[DOI no: ]
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PubMed id
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Abstract
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Major urinary proteins (MUPs) form an ensemble of protein isoforms which are
expressed and secreted by sexually mature male mice only. They belong to the
lipocalin superfamily and share with other members of this family the capacity
to bind hydrophobic molecules, some of which are odorants. MUPs, either
associated with or free of their natural ligands, play an important role in the
reproductive cycle of these rodents by acting as pheromones. In fact, they are
able to interact with receptors in the vomeronasal organ of the female mice,
inducing hormonal and physiological responses by an as yet unknown mechanism. In
order to investigate the structural and dynamical features of these proteins in
solution, one of the various wild-type isoforms (rMUP: 162 residues) was cloned
and subsequently isotopically labeled. The complete 1H, 13C and 15N resonance
assignment of that isoform, achieved by using a variety of multidimensional
heteronuclear NMR experiments, has been reported recently. Here, we describe the
refined high-resolution three-dimensional solution structure of rMUP in the
native state, obtained by a combination of distance geometry and energy
minimization calculations based on 2362 NOE-derived distance restraints. A
comparison with the crystal structure of the wild-type MUPs reveals, aside from
minor differences, a close resemblance in both secondary structure and overall
topology. The secondary structure of the protein consists of eight antiparallel
beta-strands forming a single beta-sheet and an alpha-helix in the C-terminal
region. In addition, there are several helical and hairpin turns distributed
throughout the protein sequence, mostly connecting the beta-strands. The
tertiary fold of the beta-sheet creates a beta-barrel, common to all members of
the lipocalin superfamily. The shape of the beta-barrel resembles a calyx, lined
inside by mostly hydrophobic residues that are instrumental for the binding and
transport of small nonpolar ligand molecules.
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Figure 5.
Fig. 5 Backbone drawing of the solution 3D structure of
rMUP. This figure represents the lowest energy structure
obtained for rMUP in solution. The antiparallel  -strands,
labeled A through H, form a -barrel
structure that defines the hydrophobic pocket in the protein
interior. (Produced with MOLSCRIPT [42].)
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Figure 6.
Fig. 6 Graphical representation of the hydrophobic
binding pocket inside rMUP. The side-chains of all residues in
the interior of the -barrel are
displayed as rods. The nonpolar side-chains are colored in green
(Trp19) and yellow, while polar residues are shown in red.
(Produced with GRASP [43].)
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The above figures are
reprinted
by permission from the Federation of European Biochemical Societies:
Eur J Biochem
(1999,
266,
1210-1218)
copyright 1999.
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Secondary reference #1
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Title
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Complete 1h, 15n and 13c assignment of a recombinant mouse major urinary protein.
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Authors
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F.Abbate,
L.Franzoni,
F.Löhr,
C.Lücke,
E.Ferrari,
R.T.Sorbi,
H.Rüterjans,
A.Spisni.
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Ref.
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J Biomol Nmr, 1999,
15,
187-188.
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PubMed id
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Secondary reference #2
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Title
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Expression of a lipocalin in pichia pastoris: secretion, Purification and binding activity of a recombinant mouse major urinary protein.
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Authors
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E.Ferrari,
T.Lodi,
R.T.Sorbi,
R.Tirindelli,
A.Cavaggioni,
A.Spisni.
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Ref.
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Febs Lett, 1997,
401,
73-77.
[DOI no: ]
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PubMed id
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Figure 3.
Fig. 3. A: PCR amplifications of the genomic region between
5′ and 3′ TT AOX1 sequences in six representative
transformants, MN1, MN2, MN3, MN4, MN5, MN6. Lane 6 shows the
amplification positive control, i.e. the 744 bp PCR product
(including the MUP coding segment) of the region limited by the
same AOX1 sequences in the pHIL-D2MUP vector. The MUP cDNA is
integrated in all tested Mut^s recombinants. The wild-type AOX1
gene (2.2 kb) is absent because replaced by the MUP expression
cassette. B: SDS-PAGE analysis indicating that rMUP is
accumulated in the expression culture of clone MN2 during an
induction cycle: 15 μl of medium, collected at the indicated
induction times, was loaded on the gel. Staining with Coomassie
blue detected rMUP only.
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Figure 4.
Fig. 4. A: Chromatogram of rMUP separation from the
supernatant by ion exhange chromatography. The expression
supernatant was chromatographed on a Source Q column and rMUP
was eluted with a NaCl gradient (dashed line) in Tris-HCl 10 mM
pH 7.2. The arrow indicates the rMUP elution peak. B: Binding
isotherm (18°C) of the odorant IBMP for rMUP. X-axis: free IBMP
concentration. Y-axis: bound IBMP concentration. Scatchard
analysis (inset) indicates a single set of binding sites of
K[m]=0.94 μM and a B[max]=0.88.
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The above figures are
reproduced from the cited reference
with permission from the Federation of European Biochemical Societies
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