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Biochem J. Sep 1, 2000; 350(Pt 2): 369–379.
PMCID: PMC1221263

Cloning, post-translational modifications, heterologous expression and ligand-binding of boar salivary lipocalin.


Boar submaxillary glands produce the sex-specific salivary lipocalin (SAL), which binds steroidal sex pheromones as endogenous ligands. The cDNA encoding SAL was cloned and sequenced. From a single individual, two protein isoforms, differing in three amino acid residues, were purified and structurally characterized by a combined Edman degradation/MS approach. These experiments ascertained that the mature polypeptide is composed of 168 amino acid residues, that one of the three putative glycosylation sites is post-translationally modified and the structure of the bound glycosidic moieties. Two of the cysteine residues are paired together in a disulphide bridge, whereas the remaining two occur as free thiols. SAL bears sequence similarity to other lipocalins; on this basis, a three-dimensional model of the protein has been built. A SAL isoform was expressed in Escherichia coli in good yields. Protein chemistry and CD experiments verified that the recombinant product shows the same redox state at the cysteine residues and that the same conformation is observed as in the natural protein, thus suggesting similar folding. Binding experiments on natural and recombinant SAL were performed with the fluorescent probe 1-aminoanthracene, which was efficiently displaced by the steroidal sex pheromone, as well as by several odorants.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Cavaggioni A, Findlay JB, Tirindelli R. Ligand binding characteristics of homologous rat and mouse urinary proteins and pyrazine-binding protein of calf. Comp Biochem Physiol B. 1990;96(3):513–520. [PubMed]
  • Singer AG, Macrides F, Clancy AN, Agosta WC. Purification and analysis of a proteinaceous aphrodisiac pheromone from hamster vaginal discharge. J Biol Chem. 1986 Oct 5;261(28):13323–13326. [PubMed]
  • Marchese S, Pes D, Scaloni A, Carbone V, Pelosi P. Lipocalins of boar salivary glands binding odours and pheromones. Eur J Biochem. 1998 Mar 15;252(3):563–568. [PubMed]
  • Flower DR. The lipocalin protein family: structure and function. Biochem J. 1996 Aug 15;318(Pt 1):1–14. [PMC free article] [PubMed]
  • Pelosi P. Odorant-binding proteins. Crit Rev Biochem Mol Biol. 1994;29(3):199–228. [PubMed]
  • Pelosi P. Perireceptor events in olfaction. J Neurobiol. 1996 May;30(1):3–19. [PubMed]
  • Finlayson JS, Asofsky R, Potter M, Runner CC. Major urinary protein complex of normal mice: origin. Science. 1965 Aug 27;149(3687):981–982. [PubMed]
  • Dinh BL, Tremblay A, Dufour D. Immunochemical study on rat urinary proteins: their relation to serum and kidney proteins (chromatographic separation of the major urinary protein). J Immunol. 1965 Sep;95(3):574–582. [PubMed]
  • Bacchini A, Gaetani E, Cavaggioni A. Pheromone binding proteins of the mouse, Mus musculus. Experientia. 1992 Apr 15;48(4):419–421. [PubMed]
  • Jemiolo B, Xie TM, Novotny M. Urine marking in male mice: responses to natural and synthetic chemosignals. Physiol Behav. 1992 Sep;52(3):521–526. [PubMed]
  • Henzel WJ, Rodriguez H, Singer AG, Stults JT, Macrides F, Agosta WC, Niall H. The primary structure of aphrodisin. J Biol Chem. 1988 Nov 15;263(32):16682–16687. [PubMed]
  • Shaw PH, Held WA, Hastie ND. The gene family for major urinary proteins: expression in several secretory tissues of the mouse. Cell. 1983 Mar;32(3):755–761. [PubMed]
  • Shahan K, Denaro M, Gilmartin M, Shi Y, Derman E. Expression of six mouse major urinary protein genes in the mammary, parotid, sublingual, submaxillary, and lachrymal glands and in the liver. Mol Cell Biol. 1987 May;7(5):1947–1954. [PMC free article] [PubMed]
  • Spielman AI, Zeng XN, Leyden JJ, Preti G. Proteinaceous precursors of human axillary odor: isolation of two novel odor-binding proteins. Experientia. 1995 Jan 15;51(1):40–47. [PubMed]
  • Zeng C, Spielman AI, Vowels BR, Leyden JJ, Biemann K, Preti G. A human axillary odorant is carried by apolipoprotein D. Proc Natl Acad Sci U S A. 1996 Jun 25;93(13):6626–6630. [PMC free article] [PubMed]
  • Pes D, Pelosi P. Odorant-binding proteins of the mouse. Comp Biochem Physiol B Biochem Mol Biol. 1995 Nov;112(3):471–479. [PubMed]
  • Utsumi M, Ohno K, Kawasaki Y, Tamura M, Kubo T, Tohyama M. Expression of major urinary protein genes in the nasal glands associated with general olfaction. J Neurobiol. 1999 May;39(2):227–236. [PubMed]
  • Garibotti M, Navarrini A, Pisanelli AM, Pelosi P. Three odorant-binding proteins from rabbit nasal mucosa. Chem Senses. 1997 Aug;22(4):383–390. [PubMed]
  • Ganni M, Garibotti M, Scaloni A, Pucci P, Pelosi P. Microheterogeneity of odorant-binding proteins in the porcupine revealed by N-terminal sequencing and mass spectrometry. Comp Biochem Physiol B Biochem Mol Biol. 1997 Jun;117(2):287–291. [PubMed]
  • Pes D, Mameli M, Andreini I, Krieger J, Weber M, Breer H, Pelosi P. Cloning and expression of odorant-binding proteins Ia and Ib from mouse nasal tissue. Gene. 1998 May 28;212(1):49–55. [PubMed]
  • Mucignat-Caretta C, Caretta A, Cavaggioni A. Acceleration of puberty onset in female mice by male urinary proteins. J Physiol. 1995 Jul 15;486(Pt 2):517–522. [PMC free article] [PubMed]
  • Krieger J, Schmitt A, Löbel D, Gudermann T, Schultz G, Breer H, Boekhoff I. Selective activation of G protein subtypes in the vomeronasal organ upon stimulation with urine-derived compounds. J Biol Chem. 1999 Feb 19;274(8):4655–4662. [PubMed]
  • Ryba NJ, Tirindelli R. A new multigene family of putative pheromone receptors. Neuron. 1997 Aug;19(2):371–379. [PubMed]
  • Herrada G, Dulac C. A novel family of putative pheromone receptors in mammals with a topographically organized and sexually dimorphic distribution. Cell. 1997 Aug 22;90(4):763–773. [PubMed]
  • Matsunami H, Buck LB. A multigene family encoding a diverse array of putative pheromone receptors in mammals. Cell. 1997 Aug 22;90(4):775–784. [PubMed]
  • Katkov T, Booth WD, Gower DB. The metabolism of 16-androstenes in boar salivary glands. Biochim Biophys Acta. 1972 Aug 11;270(4):546–556. [PubMed]
  • Booth WD, White CA. The isolation, purification and some properties of pheromaxein, the pheromonal steroid-binding protein, in porcine submaxillary glands and saliva. J Endocrinol. 1988 Jul;118(1):47–57. [PubMed]
  • Booth WD, von Glos KI. Pheromaxein, the pheromonal steroid-binding protein, is a major protein synthesized in porcine submaxillary salivary glands. J Endocrinol. 1991 Feb;128(2):205–212. [PubMed]
  • Freitag J, Beck A, Ludwig G, von Buchholtz L, Breer H. On the origin of the olfactory receptor family: receptor genes of the jawless fish (Lampetra fluviatilis). Gene. 1999 Jan 21;226(2):165–174. [PubMed]
  • Engler-Blum G, Meier M, Frank J, Müller GA. Reduction of background problems in nonradioactive northern and Southern blot analyses enables higher sensitivity than 32P-based hybridizations. Anal Biochem. 1993 May 1;210(2):235–244. [PubMed]
  • Löbel D, Marchese S, Krieger J, Pelosi P, Breer H. Subtypes of odorant-binding proteins--heterologous expression and ligand binding. Eur J Biochem. 1998 Jun 1;254(2):318–324. [PubMed]
  • Kyhse-Andersen J. Electroblotting of multiple gels: a simple apparatus without buffer tank for rapid transfer of proteins from polyacrylamide to nitrocellulose. J Biochem Biophys Methods. 1984 Dec;10(3-4):203–209. [PubMed]
  • Angeli S, Ceron F, Scaloni A, Monti M, Monteforti G, Minnocci A, Petacchi R, Pelosi P. Purification, structural characterization, cloning and immunocytochemical localization of chemoreception proteins from Schistocerca gregaria. Eur J Biochem. 1999 Jun;262(3):745–754. [PubMed]
  • Thompson JD, Higgins DG, Gibson TJ. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994 Nov 11;22(22):4673–4680. [PMC free article] [PubMed]
  • Böcskei Z, Groom CR, Flower DR, Wright CE, Phillips SE, Cavaggioni A, Findlay JB, North AC. Pheromone binding to two rodent urinary proteins revealed by X-ray crystallography. Nature. 1992 Nov 12;360(6400):186–188. [PubMed]
  • Spinelli S, Ramoni R, Grolli S, Bonicel J, Cambillau C, Tegoni M. The structure of the monomeric porcine odorant binding protein sheds light on the domain swapping mechanism. Biochemistry. 1998 Jun 2;37(22):7913–7918. [PubMed]
  • Bianchet MA, Bains G, Pelosi P, Pevsner J, Snyder SH, Monaco HL, Amzel LM. The three-dimensional structure of bovine odorant binding protein and its mechanism of odor recognition. Nat Struct Biol. 1996 Nov;3(11):934–939. [PubMed]
  • Tegoni M, Ramoni R, Bignetti E, Spinelli S, Cambillau C. Domain swapping creates a third putative combining site in bovine odorant binding protein dimer. Nat Struct Biol. 1996 Oct;3(10):863–867. [PubMed]
  • Peitsch MC. ProMod and Swiss-Model: Internet-based tools for automated comparative protein modelling. Biochem Soc Trans. 1996 Feb;24(1):274–279. [PubMed]
  • Andrade MA, Chacón P, Merelo JJ, Morán F. Evaluation of secondary structure of proteins from UV circular dichroism spectra using an unsupervised learning neural network. Protein Eng. 1993 Jun;6(4):383–390. [PubMed]
  • Paolini S, Tanfani F, Fini C, Bertoli E, Paolo Pelosi Porcine odorant-binding protein: structural stability and ligand affinities measured by fourier-transform infrared spectroscopy and fluorescence spectroscopy. Biochim Biophys Acta. 1999 Apr 12;1431(1):179–188. [PubMed]
  • Gregoire C, Rosinski-Chupin I, Rabillon J, Alzari PM, David B, Dandeu JP. cDNA cloning and sequencing reveal the major horse allergen Equ c1 to be a glycoprotein member of the lipocalin superfamily. J Biol Chem. 1996 Dec 20;271(51):32951–32959. [PubMed]
  • Gao F, Endo H, Yamamoto M. Length heterogeneity in rat salivary gland alpha 2 mu globulin mRNAs: multiple splice-acceptors and polyadenylation sites. Nucleic Acids Res. 1989 Jun 26;17(12):4629–4636. [PMC free article] [PubMed]

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