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Glycation Contributes to Interaction Between Human Bone Alkaline Phosphatase and Collagen Type I
Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences. Östergötlands Läns Landsting, Center for Diagnostics, Department of Clinical Chemistry.
Linköping University, Department of Physics, Chemistry and Biology, Molecular Physics. Linköping University, Faculty of Science & Engineering.
Linköping University, Department of Clinical and Experimental Medicine, Division of Microbiology and Molecular Medicine. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Center for Diagnostics, Department of Clinical Chemistry.
2016 (English)In: Calcified Tissue International, ISSN 0171-967X, E-ISSN 1432-0827, Vol. 98, no 3, 284-293 p.Article in journal (Refereed) PublishedText
Abstract [en]

Bone is a biological composite material comprised primarily of collagen type I and mineral crystals of calcium and phosphate in the form of hydroxyapatite (HA), which together provide its mechanical properties. Bone alkaline phosphatase (ALP), produced by osteoblasts, plays a pivotal role in the mineralization process. Affinity contacts between collagen, mainly type II, and the crown domain of various ALP isozymes were reported in a few in vitro studies in the 1980s and 1990s, but have not attracted much attention since, although such interactions may have important implications for the bone mineralization process. The objective of this study was to investigate the binding properties of human collagen type I to human bone ALP, including the two bone ALP isoforms B1 and B2. ALP from human liver, human placenta and E. coli were also studied. A surface plasmon resonance-based analysis, supported by electrophoresis and blotting, showed that bone ALP binds stronger to collagen type I in comparison with ALPs expressed in non-mineralizing tissues. Further, the B2 isoform binds significantly stronger to collagen type I in comparison with the B1 isoform. Human bone and liver ALP (with identical amino acid composition) displayed pronounced differences in binding, revealing that post-translational glycosylation properties govern these interactions to a large extent. In conclusion, this study presents the first evidence that glycosylation differences in human ALPs are of crucial importance for protein–protein interactions with collagen type I, although the presence of the ALP crown domain may also be necessary. Different binding affinities among the bone ALP isoforms may influence the mineral-collagen interface, mineralization kinetics, and degree of bone matrix mineralization, which are important factors determining the material properties of bone.

Place, publisher, year, edition, pages
Springer-Verlag New York, 2016. Vol. 98, no 3, 284-293 p.
Keyword [en]
Alkaline phosphatase; Bone; Collagen; Glycosylation; Mineralization; Surface plasmon resonance
National Category
Endocrinology and Diabetes Dentistry
Identifiers
URN: urn:nbn:se:liu:diva-127099DOI: 10.1007/s00223-015-0088-0ISI: 000373744700008PubMedID: 26645431OAI: oai:DiVA.org:liu-127099DiVA: diva2:919469
Note

Funding agencies:  Region Ostergotland, Sweden

Available from: 2016-04-14 Created: 2016-04-14 Last updated: 2016-05-04Bibliographically approved
In thesis
1. Biochemical and functional properties of mammalian bone alkaline phosphatase isoforms during osteogenesis
Open this publication in new window or tab >>Biochemical and functional properties of mammalian bone alkaline phosphatase isoforms during osteogenesis
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The human skeleton is a living and dynamic tissue that constantly is being renewed in a process called bone remodeling. Old bone is resorbed by osteoclasts and new bone is formed by osteoblasts. Bone is a composite material made up by mineral crystals in the form of hydroxyapatite (calcium and phosphate) that provides the hardness of bone, and collagen fibrils that provides elasticity and flexibility. Alkaline phosphatase (ALP) is a family of enzymes that is present in most species and catalyzes the hydrolysis of various phosphomonoesters at alkaline pH. Despite the generalized use of ALP as a biochemical marker of bone formation, the precise function of bone ALP (BALP) is only now becoming clear. Three circulating human BALP isoforms (B1, B2, and B/I) can be distinguished in healthy individuals and a fourth isoform (B1x) has been discovered in patients with chronic kidney disease and in bone tissue.

Paper I. Three endogenous phosphocompounds, (i.e., inorganic pyrophosphate (PPi), pyridoxal 5′-phosphate (PLP) and phosphoethanolamine (PEA)), have been suggested to serve as  physiological substrates for BALP. The BALP isoforms display different catalytic properties towards PPi and PLP, which is attributed to their distinct N-linked glycosylation patterns. The catalytic activity, using PEA as substrate, was barely detectable for all BALP isoforms indicating that PEA is not a physiological substrate for BALP.

Paper II. Mouse serum ALP is frequently measured and interpreted in mammalian bone research. However, little is known about the circulating ALPs in mice and their relation to human ALP. We characterized the circulating and tissue-derived mouse ALP isozymes and isoforms from mixed strains of wild-type and knockout mice. All four BALP isoforms (B/I, B1x, B1, and B2) were identified in mouse serum and bone tissues, in good correspondence with those found in human bones. All mouse tissues, except liver, contained significant ALP activities. This is a notable difference as human liver contains vast amounts of ALP.

Paper III. The objective of this study was to investigate the binding properties of human collagen type I to human BALP, including the two BALP isoforms B1 and B2, together with ALP from human liver, human placenta and E. coli. A surface plasmon resonance-based analysis showed that BALP binds stronger to collagen type I in comparison with ALPs expressed in non-mineralizing tissues. The B2 isoform binds significantly stronger to collagen type I in comparison with the B1 isoform, indicating that glycosylation differences in human ALPs are of crucial importance for protein–protein interactions with collagen type I.

Paper IV. Tartrate-resistant acid phosphatase (TRAP) is highly expressed in osteoclasts and frequently used as a marker of bone resorption. Intriguingly, recent studies show that TRAP is also expressed in osteoblasts and osteocytes. TRAP displays enzymatic activity towards the endogenous substrates for BALP, i.e., PPi and PLP. Both TRAP and BALP can alleviate the inhibitory effect of osteopontin on mineralization by dephosphorylation, which suggests a novel role for TRAP in skeletal mineralization.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2016. 66 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1511
National Category
Clinical Medicine Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Biomaterials Science Cell and Molecular Biology
Identifiers
urn:nbn:se:liu:diva-127100 (URN)10.3384/diss.diva-127100 (DOI)978-91-7685-824-0 (Print) (ISBN)
Public defence
2016-05-13, Berzeliussalen, Campus Valla, Linköping, 09:00 (Swedish)
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Available from: 2016-04-14 Created: 2016-04-14 Last updated: 2016-04-14Bibliographically approved

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