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Structural / functi...
Structural / functional studies on human alkaline phosphatases
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- Kozlenkov, Alexey, 1973- (author)
- Umeå universitet,Institutionen för medicinsk biovetenskap
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Millan, Jose-Luis (thesis advisor)
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- Jörnvall, Hans, Professor (opponent)
- Institutionen för medisinsk biokemi och biofysik, Fysiologisk kemi, Karolinska insitutet, Stockholm
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(creator_code:org_t)
- ISBN 9173055425
- 2003
- English 128 s.
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Series: Umeå University medical dissertations, 0346-6612 ; 863
- Related links:
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https://urn.kb.se/re...
Abstract
Subject headings
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- The recent elucidation of the three-dimensional structure of human placental alkaline phosphatase (PLAP) has enabled me to perform structural studies aimed at characterizing the properties of human PLAP and tissue-nonspecific AP (TNAP) as paradigms for mammalian APs in general, using site-directed mutagenesis, protein expression, kinetic analysis and computer modeling. In Paper I, we found that a single critical E429G substitution explains the difference in stability and kinetics between the common allelic variants of PLAP and the D allozyme. In Paper II, we demonstrated the role of residue E429 in PLAP in stabilizing the active site metals, elucidated the distinct roles of residues H153 and H317 in catalysis, and the relative importance of five Cys residues in PLAP. We also discovered the significance of Y367, a unique feature of mammalian APs, for enzyme stability and specific inhibition by amino acids. Paper III focused on the identification and mutagenesis analysis of a novel, non-catalytic peripheral binding site of PLAP that appears to mediate a mitogenic effect of PLAP. This site provides indications that PLAP may function as a fetal growth factor. The last two papers focus on the TNAP isozyme as paradigm. A deficiency in TNAP activity is the cause of the human disease hypophosphatasia, characterized by rickets, osteomalacia and occasionally epileptic seizures. Paper IV has been able to partially explain the variable expressivity of hypophosphatasia traits by examining site-directed mutants of TNAP and performing kinetic analysis using natural substrates PPi and PLP. Finally, Paper V has clarified the mechanism of inhibition of TNAP by uncompetitive inhibitors L-homoarginine, levamisole and theophylline. We identified residues that confer to TNAP its distinct inhibitory properties. These data have significance for future drug design of specific TNAP inhibitors to therapeutically target TNAP as a way of elevating PPi extracellular level and alleviating pathological bone hypermineralization conditions.
Subject headings
- MEDICIN OCH HÄLSOVETENSKAP -- Medicinska och farmaceutiska grundvetenskaper -- Mikrobiologi inom det medicinska området (hsv//swe)
- MEDICAL AND HEALTH SCIENCES -- Basic Medicine -- Microbiology in the medical area (hsv//eng)
Keyword
- Biomedicine
- mutagenesis
- enzyme kinetics
- computer modeling
- crystallography
- natural substrates
- uncompetitive inhibition
- isozymes
- gene families
- Biomedicin
- Microbiology
- Mikrobiologi
- medicinsk genetik
- Medical Genetics
Publication and Content Type
- vet (subject category)
- dok (subject category)
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