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. 1999 Dec;8(12):2611–2620. doi: 10.1110/ps.8.12.2611

Alpha1-microglobulin chromophores are located to three lysine residues semiburied in the lipocalin pocket and associated with a novel lipophilic compound.

T Berggård 1, A Cohen 1, P Persson 1, A Lindqvist 1, T Cedervall 1, M Silow 1, I B Thøgersen 1, J A Jönsson 1, J J Enghild 1, B Akerström 1
PMCID: PMC2144230  PMID: 10631976

Abstract

Alpha1-microglobulin (alpha1m) is an electrophoretically heterogeneous plasma protein. It belongs to the lipocalin superfamily, a group of proteins with a three-dimensional (3D) structure that forms an internal hydrophobic ligand-binding pocket. Alpha1m carries a covalently linked unidentified chromophore that gives the protein a characteristic brown color and extremely heterogeneous optical properties. Twenty-one different colored tryptic peptides corresponding to residues 88-94, 118-121, and 122-134 of human alpha1m were purified. In these peptides, the side chains of Lys92, Lys118, and Lys130 carried size heterogeneous, covalently attached, unidentified chromophores with molecular masses between 122 and 282 atomic mass units (amu). In addition, a previously unknown uncolored lipophilic 282 amu compound was found strongly, but noncovalently associated with the colored peptides. Uncolored tryptic peptides containing the same Lys residues were also purified. These peptides did not carry any additional mass (i.e., chromophore) suggesting that only a fraction of the Lys92, Lys118, and Lys130 are modified. The results can explain the size, charge, and optical heterogeneity of alpha1m. A 3D model of alpha1m, based on the structure of rat epididymal retinoic acid-binding protein (ERABP), suggests that Lys92, Lys118, and Lys130 are semiburied near the entrance of the lipocalin pocket. This was supported by the fluorescence spectra of alpha1m under native and denatured conditions, which indicated that the chromophores are buried, or semiburied, in the interior of the protein. In human plasma, approximately 50% of alpha1m is complex bound to IgA. Only the free alpha1m carried colored groups, whereas alpha1m linked to IgA was uncolored.

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Selected References

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  1. Akerström B., Bratt T., Enghild J. J. Formation of the alpha 1-microglobulin chromophore in mammalian and insect cells: a novel post-translational mechanism? FEBS Lett. 1995 Mar 27;362(1):50–54. doi: 10.1016/0014-5793(95)00206-o. [DOI] [PubMed] [Google Scholar]
  2. Akerström B., Lögdberg L. An intriguing member of the lipocalin protein family: alpha 1-microglobulin. Trends Biochem Sci. 1990 Jun;15(6):240–243. doi: 10.1016/0968-0004(90)90037-c. [DOI] [PubMed] [Google Scholar]
  3. Babiker-Mohamed H., Forsberg M., Olsson M. L., Winquist O., Nilson B. H., Lögdberg L., Akerström B. Characterization of monoclonal anti-alpha 1-microglobulin antibodies: binding strength, binding sites, and inhibition of lymphocyte stimulation. Scand J Immunol. 1991 Nov;34(5):655–666. doi: 10.1111/j.1365-3083.1991.tb01589.x. [DOI] [PubMed] [Google Scholar]
  4. Berggård T., Thelin N., Falkenberg C., Enghild J. J., Akerström B. Prothrombin, albumin and immunoglobulin A form covalent complexes with alpha1-microglobulin in human plasma. Eur J Biochem. 1997 May 1;245(3):676–683. doi: 10.1111/j.1432-1033.1997.00676.x. [DOI] [PubMed] [Google Scholar]
  5. Bonay P., Solís J., de la Calle H., Fresno M., Grubb A., Méndez E. Massive glycation of protein HC, a low molecular weight lipocalin, in non-diabetic individuals. FEBS Lett. 1997 Oct 27;416(3):276–280. doi: 10.1016/s0014-5793(97)01220-9. [DOI] [PubMed] [Google Scholar]
  6. Bratt T., Olsson H., Sjöberg E. M., Jergil B., Akerström B. Cleavage of the alpha 1-microglobulin-bikunin precursor is localized to the Golgi apparatus of rat liver cells. Biochim Biophys Acta. 1993 Jun 11;1157(2):147–154. doi: 10.1016/0304-4165(93)90058-g. [DOI] [PubMed] [Google Scholar]
  7. Calero M., Escribano J., Grubb A., Méndez E. Location of a novel type of interpolypeptide chain linkage in the human protein HC-IgA complex (HC-IgA) and identification of a heterogeneous chromophore associated with the complex. J Biol Chem. 1994 Jan 7;269(1):384–389. [PubMed] [Google Scholar]
  8. Chappey O., Dosquet C., Wautier M. P., Wautier J. L. Advanced glycation end products, oxidant stress and vascular lesions. Eur J Clin Invest. 1997 Feb;27(2):97–108. doi: 10.1046/j.1365-2362.1997.710624.x. [DOI] [PubMed] [Google Scholar]
  9. Deyl Z., Miksík I. Post-translational non-enzymatic modification of proteins. I. Chromatography of marker adducts with special emphasis to glycation reactions. J Chromatogr B Biomed Sci Appl. 1997 Oct 10;699(1-2):287–309. doi: 10.1016/s0378-4347(97)00163-1. [DOI] [PubMed] [Google Scholar]
  10. Ekström B., Berggård I. Human alpha1-microglobulin. Purification procedure, chemical and physiochemical properties. J Biol Chem. 1977 Nov 25;252(22):8048–8057. [PubMed] [Google Scholar]
  11. Ekström B., Peterson P. A., Berggård I. A urinary and plasma alpha1-glycoprotein of low molecular weight: isolation and some properties. Biochem Biophys Res Commun. 1975 Aug 18;65(4):1427–1433. doi: 10.1016/s0006-291x(75)80388-3. [DOI] [PubMed] [Google Scholar]
  12. Enghild J. J., Thøgersen I. B., Pizzo S. V., Salvesen G. Analysis of inter-alpha-trypsin inhibitor and a novel trypsin inhibitor, pre-alpha-trypsin inhibitor, from human plasma. Polypeptide chain stoichiometry and assembly by glycan. J Biol Chem. 1989 Sep 25;264(27):15975–15981. [PubMed] [Google Scholar]
  13. Escribano J., Grubb A., Calero M., Méndez E. The protein HC chromophore is linked to the cysteine residue at position 34 of the polypeptide chain by a reduction-resistant bond and causes the charge heterogeneity of protein HC. J Biol Chem. 1991 Aug 25;266(24):15758–15763. [PubMed] [Google Scholar]
  14. Escribano J., Lopex-Otin C., Hjerpe A., Grubb A., Mendez E. Location and characterization of the three carbohydrate prosthetic groups of human protein HC. FEBS Lett. 1990 Jun 18;266(1-2):167–170. doi: 10.1016/0014-5793(90)81531-r. [DOI] [PubMed] [Google Scholar]
  15. Falkenberg C., Enghild J. J., Thøgersen I. B., Salvesen G., Akerström B. Isolation and characterization of fibronectin-alpha 1-microglobulin complex in rat plasma. Biochem J. 1994 Aug 1;301(Pt 3):745–751. doi: 10.1042/bj3010745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Falkenberg C., Grubb A., Akerström B. Isolation of rat serum alpha 1-microglobulin. Identification of a complex with alpha 1-inhibitor-3, a rat alpha 2-macroglobulin homologue. J Biol Chem. 1990 Sep 25;265(27):16150–16157. [PubMed] [Google Scholar]
  17. Flower D. R. Multiple molecular recognition properties of the lipocalin protein family. J Mol Recognit. 1995 May-Jun;8(3):185–195. doi: 10.1002/jmr.300080304. [DOI] [PubMed] [Google Scholar]
  18. Flower D. R. The lipocalin protein family: structure and function. Biochem J. 1996 Aug 15;318(Pt 1):1–14. doi: 10.1042/bj3180001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Grubb A. O., López C., Tejler L., Mendez E. Isolation of human complex-forming glycoprotein, heterogeneous in charge (protein HC), and its IgA complex from plasma. Physiochemical and immunochemical properties, normal plasma concentration. J Biol Chem. 1983 Dec 10;258(23):14698–14707. [PubMed] [Google Scholar]
  20. Guex N., Peitsch M. C. SWISS-MODEL and the Swiss-PdbViewer: an environment for comparative protein modeling. Electrophoresis. 1997 Dec;18(15):2714–2723. doi: 10.1002/elps.1150181505. [DOI] [PubMed] [Google Scholar]
  21. Hanley S., McInerney J. O., Powell R. Sequence analysis and evolutionary aspects of piscine alpha-1-microglobulin/bikunin mRNA transcripts. Mol Mar Biol Biotechnol. 1995 Jun;4(2):105–109. [PubMed] [Google Scholar]
  22. Hanley S., Powell R. Sequence of a cDNA clone encoding the Atlantic salmon alpha 1-microglobulin/bikunin protein. Gene. 1994 Sep 30;147(2):297–298. doi: 10.1016/0378-1119(94)90086-8. [DOI] [PubMed] [Google Scholar]
  23. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  24. Leaver M. J., Wright J., George S. G. Conservation of the tandem arrangement of alpha 1-microglobulin/bikunin mRNA: cloning of a cDNA from plaice (Pleuronectes platessa). Comp Biochem Physiol Biochem Mol Biol. 1994 Jul;108(3):275–281. doi: 10.1016/0305-0491(94)90077-9. [DOI] [PubMed] [Google Scholar]
  25. Lindqvist A., Akerström B. Bovine alpha 1-microglobulin/bikunin. Isolation and characterization of liver cDNA and urinary alpha 1-microglobulin. Biochim Biophys Acta. 1996 Apr 10;1306(1):98–106. doi: 10.1016/0167-4781(95)00235-9. [DOI] [PubMed] [Google Scholar]
  26. Lindqvist A., Akerström B. Isolation of plaice (Pleuronectes platessa) alpha1-microglobulin: conservation of structure and chromophore. Biochim Biophys Acta. 1999 Mar 19;1430(2):222–233. doi: 10.1016/s0167-4838(99)00003-5. [DOI] [PubMed] [Google Scholar]
  27. Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
  28. Salier J. P. Inter-alpha-trypsin inhibitor: emergence of a family within the Kunitz-type protease inhibitor superfamily. Trends Biochem Sci. 1990 Nov;15(11):435–439. doi: 10.1016/0968-0004(90)90282-g. [DOI] [PubMed] [Google Scholar]
  29. Sansom C. E., North A. C., Sawyer L. Structural analysis and classification of lipocalins and related proteins using a profile-search method. Biochim Biophys Acta. 1994 Oct 19;1208(2):247–255. doi: 10.1016/0167-4838(94)90110-4. [DOI] [PubMed] [Google Scholar]
  30. Tejler L., Grubb A. O. A complex-forming glycoprotein heterogeneous in charge and present in human plasma, urine, and cerebrospinal fluid. Biochim Biophys Acta. 1976 Jul 19;439(1):82–94. doi: 10.1016/0005-2795(76)90164-1. [DOI] [PubMed] [Google Scholar]
  31. Thorpe S. R., Baynes J. W. Role of the Maillard reaction in diabetes mellitus and diseases of aging. Drugs Aging. 1996 Aug;9(2):69–77. doi: 10.2165/00002512-199609020-00001. [DOI] [PubMed] [Google Scholar]
  32. Thøgersen I. B., Enghild J. J. Biosynthesis of bikunin proteins in the human carcinoma cell line HepG2 and in primary human hepatocytes. Polypeptide assembly by glycosaminoglycan. J Biol Chem. 1995 Aug 4;270(31):18700–18709. doi: 10.1074/jbc.270.31.18700. [DOI] [PubMed] [Google Scholar]
  33. Wester L., Johansson M. U., Akerström B. Physicochemical and biochemical characterization of human alpha 1-microglobulin expressed in baculovirus-infected insect cells. Protein Expr Purif. 1997 Oct;11(1):95–103. doi: 10.1006/prep.1997.0760. [DOI] [PubMed] [Google Scholar]

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