A1M: a heme and radical binding protein. A study on structure, function and mechanisms
Författare
Summary, in English
This thesis describes functional and structural studies of the human protein α1-microglobulin (A1M), an evolutionarily well-conserved 26 kDa plasma and tissue protein mainly synthesized in the liver. A1M belongs to the lipocalin protein family. Lipocalins share a very similar structur, an 8-stranded β-barrel forming a hydrophobic pocket. A1M has reductase and radical scavenging properties. A1M also binds the iron-containing heme-group, a lipophilic small compound found in many proteins and enzymes, but toxic in free form. A truncated form of A1M (t-A1M) catalyzes degradation of the heme-group. These properties give A1M antioxidative properties in human tissues.
The research focus in this thesis has been to study the protection mechanisms of A1M with a special focus on the structure, and to investigate its protection properties in vivo.
The results show that radiation-induced cell damage induces reactive oxygen species (ROS) and that ROS mediates a so-called bystander effect (a spreading of damage to cells not directly hit by the radiation). A1M inhibited the bystander effect and by using site-directed mutagenesis it was shown that this protection is dependent on a cysteine side-chain in position 34, and regulated by three lysine residues in position 92, 118 and 130.
Furthermore, when A1M binds heme, the cysteine in position 34, a histidine in position 123, and the three lysine-residues in position 92, 118 and 130 are involved in co-ordination of the iron-atom in heme. The results support a model in which two heme-groups are bound simultaneously. Molecular simulation suggests binding of the first heme group inside the lipocalin pocket, followed by a structural shift that allows binding of a second heme group, with lower affinity, near the opening of pocket.
Finally, the in vivo therapeutic effects of A1M were investigated using a sheep model of the pregnancy disease preeclampsia. Starvation induced preeclampsia-like symptoms in the pregnant ewes manifesting in hemolysis and subsequent damage to the placenta and kidneys. A1M was well tolerated and displayed positive therapeutic effects.
The research focus in this thesis has been to study the protection mechanisms of A1M with a special focus on the structure, and to investigate its protection properties in vivo.
The results show that radiation-induced cell damage induces reactive oxygen species (ROS) and that ROS mediates a so-called bystander effect (a spreading of damage to cells not directly hit by the radiation). A1M inhibited the bystander effect and by using site-directed mutagenesis it was shown that this protection is dependent on a cysteine side-chain in position 34, and regulated by three lysine residues in position 92, 118 and 130.
Furthermore, when A1M binds heme, the cysteine in position 34, a histidine in position 123, and the three lysine-residues in position 92, 118 and 130 are involved in co-ordination of the iron-atom in heme. The results support a model in which two heme-groups are bound simultaneously. Molecular simulation suggests binding of the first heme group inside the lipocalin pocket, followed by a structural shift that allows binding of a second heme group, with lower affinity, near the opening of pocket.
Finally, the in vivo therapeutic effects of A1M were investigated using a sheep model of the pregnancy disease preeclampsia. Starvation induced preeclampsia-like symptoms in the pregnant ewes manifesting in hemolysis and subsequent damage to the placenta and kidneys. A1M was well tolerated and displayed positive therapeutic effects.
Avdelning/ar
Publiceringsår
2014
Språk
Engelska
Publikation/Tidskrift/Serie
Lund University Faculty of Medicine Doctoral Dissertation Series
Volym
2014:139
Dokumenttyp
Doktorsavhandling
Förlag
Division of Infection Medicine
Ämne
- Infectious Medicine
Nyckelord
- α1-microglobulin
- oxidative stress
- heme
- radiation
- reactive oxygen species
- preeclampsia
Status
Published
Handledare
ISBN/ISSN/Övrigt
- ISSN: 1652-8220
- ISBN: 978-91-7619-068-5
Försvarsdatum
13 december 2014
Försvarstid
10:00
Försvarsplats
Belfrage, BMC D15, Klinikgatan 32, Lund
Opponent
- Maria D. Ganfornina (Professor)