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Institute of Physiological Chemistry
 

Research Group “Applied Molecular Biology”

The focus of the ERC Advanced Investigator Grant research group of Prof. Dr. Werner E. G. Müller, which includes the groups of Prof. Dr. Xiaohong Wang, Prof. Dr. Dr. Heinz C. Schröder and Dr. Matthias Wiens, concerns the following Topics:

Inorganic Functional Biopolymers and Molecular Biomineralization

Starting with the evolutionary oldest animals, the siliceous sponges, up to humans, the principle molecular processes underlying biomineral formation are studied. The group succeeeded to show that both the formation of the biosilica skeleton of the siliceous sponges and the deposition of the inorganic bone material (hydroxyapatite, HA) proceeds and is regulated via enzymatic mechanisms. The biocatalytic and structure-guiding functions of the silicateins (biosilica synthesis in the siliceous sponges) as well as the carbonic anhydrase-mediated formation of calcium carbonate (skeleton of the calcareous sponges) have been elucidated. These discoveries allowed the development of novel applications from biomedicine (implants) to nanotechnology/microoptics (light transmission). Moreover, we were the first to show that the mineralisation of human bone is initiated by a carbonic anhydrase (CA IX) mediated formation of „bioseeds“, consisting of amorphous calcium carbonate (ACC), that are subsequently transformed by carbonate-phosphate-exchange into amorphous calcium phosphate (ACP, HA precursor). Thereby the phosphate is provided, again enzymatically, by hydrolysis of inorganic polyphosphate (polyP) via the alkaline phosphatase (ALP). Recently we could show that the long-chain polyP molecules, linked by numerous energy-rich phosphodiester bonds, serve as an extracellular energy storage/signalling molecule – a fundamentally new principle in cellular energy metabolism.  These research activities are also in the focus of the Chinese-German Center for “Bio-inspired Materials”, coordinated by Prof. Müller together with Prof. Wang.

Figur01
New insights in the initial steps of bone formation. The mineral deposition proceeds in three phases. Phase I: Formation of ACC bioseeds mediated by CA IX. Phase II: ALP-mediated hydrolysis of polyP under formation of orthophosphate for the carbonate-phosphate transfer reaction. Phase III: ACP/HA formation.
Fig02
Discovery of the role of polyP as extracellular energy transport/signaling molecule (“metabolic fuel”) – a new foundational principle in energy metabolism. PolyP released into the bloodstream is engulfed in platelets. PolyP turns on the mTOR pathway and causes an increase in the number of mitochondria. After the release into the extracellular space polyP is hydrolyzed by the ALP to orthophosphate (building block for HA formation).

Bio-inspired Materials for Regenerative Medicine

Building upon on the results of basic research, our group is developing novel strategies for the fabrication of materials for medical applications, in particular implants (bone, cartilage, blood vessels, and cornea), and for 3D-printing / bioplotting. The main focus are morphogeneticaly active inorganic polymers such as energy-rich amorphous calcium-polyphosphate particles, amorphous biosilica, amorphous calcium carbonate and calcium phosphate (ACC and ACP) as well as the combination of these building blocks with bioinert materials. Techniques for the adjustable hardening of these materials are developed. This research topic, funded in the frame of two ERC Proof-of-Concept projects (“Si-Bone-PoC” and ERC Horizon 2020 “MorphoVES-PoC”) as well as of the EU FP7 project “BioScaffolds” (coordinator: W.E.G. Müller) also comprises the fabrication of nanofiber networks by electrospinning. First regeneratively active implant materials have already been tested in animal experiments and shall be introduced into clinical application. For example, we recently succeeded to develop a novel bioinspired hydrogel, composed of morphogenetically active inorganic polyP, along with two likewise biocompatible and negatively charged but bioinert natural polymers. This material can be hardened with calcium ions to yield regenerative scaffold materials that can be used for bone or cartilage and even artificial blood vessels. These biocompatible and biodegradable materials allow a fast and complete replacement by the body’s own tissue such as bone, without the need of (stem) cells or cytokine/growth factor supplementation.

Printable multicomponent system for the fabrication of hardenable implants. In the absence of calcium ions the material can be bioprinted. After addition of calcium ions, the cation binds to the polyanions under formation of fibers that organize and orient the inert biopolymers and bioactive polyP to durable implants.
Molding procedure for the fabrication of the cornea implants, based on the use of 3D printed molds (Top). The hardened cornea implant, consisting of biocompatible biomaterials, is shown (Bottom). Middle: Innovative vascular grafts (right) suitable for small diameter applica¬tions in contrast to conventional grafts (left). Right: Extruder used for the fabrication of the artificial vessels.
Fig05
3D printing of a mould for the cartilage regions in the tibia using a moulding procedure. (A) Scanning with a laser-based 3D scanner, allowing the compilation of the STL file (B) and the preparation of a casting form for the articular cartilage surface (C). (D and E) The sequential regions of the tibia and placement of the parts on top of each other. (F) Fabrication of the moulded superficial articular cartilage surface with a plastic material.

Marine Biomolecules for Biotechnology and Biomedicine

Marine sponges are an inexhaustible source of medical or biotechnological interesting proteins and bioactive compounds. As part of the EU FP7 Large - Scale Integrating Project BlueGenics, methods for the sustainable production of these compounds are being developed, particularly for therapy/prophylaxis of bone diseases (osteoporosis). Examples for compounds investigated are isoquercitrin and Ca-polyP (synergistic action on human bone formation), quinolinic acid (activator of carbonic anhydrase involved in bone formation) and retinol/Ca-polyP nanospheres (wound healing). The further focus is on molecular biology and phylogenetic analysis of the genome of aquatic invertebrates (in particular sponges) to elucidate the origin of Metazoa about 700 million years ago. Functional studies concern molecules of innate immunity, apoptosis, cell communication and morphogenesis. The analysis contributes to the understanding of basic biochemical and pathobiochemical mechanisms in humans.

Fig06
Differential modes of action of isoquercitrin and Ca-polyP, two synergistically acting molecules stimulating bone formation.
Fig07
Carbonic anhydrase (CA) involved in human bone formation as a novel drug target for CA activators. Proposed interaction of quinolinic acid (QA) with the active center of the human CA.
Fig08
Synergistic action elicited by the components (polyP and retinol) of retinol/Ca-polyP nanospheres during collagen type III gene expression.

The Research Group with its Center of Excellence “BIOTECmarin - Biomaterials from the Sea” has been selected as one of the winners in the frame of the national initiative “Germany - Land of Ideas”. The Group is intensively involved in the training of graduate and postgraduate students and has coordinated several EU projects in this field such as the Marie Curie Research Training Networks “BioCapital” (coordinator: WEG Müller) and “BIOMINTEC” (coordinator: HC Schröder), as well as the EU FP7 Industry - Academia Partnerships and Pathways project “CoreShell” (coordinator: XH Wang ) and the EU FP7 Marie Curie staff exchange project “MarBioTec” (“European - Chinese Research Staff Exchange Cluster on Marine Biotechnology”; coordinator: WEG Müller) .

Further Information about the Research Group