Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Dilute self-healing hydrogels of silk-collagen-like block copolypeptides at neutral pH
Golinska, M.D. ; Wlodarczyk-Biegun, M.K. ; Werten, M.W.T. ; Cohen Stuart, M.A. ; Wolf, F.A. de; Vries, R.J. de - \ 2014
Biomacromolecules 15 (2014)3. - ISSN 1525-7797 - p. 699 - 706.
spider dragline silk - drug-delivery - fibrous biomaterials - actin solutions - contact-lenses - scaffolds - gels - peptides - polymers - elasticity
We report on self-healing, pH-responsive hydrogels that are entirely protein-based. The protein is a denovo designed recombinant triblock polypeptide of 66 kg/mol consisting of a silk-like middle block (GAGAGAGH)48, flanked by two long collagen-inspired hydrophilic random coil side blocks. The pH-dependent charge on the histidines in the silk block controls folding and stacking of the silk block. At low pH the protein exists as monomers, but above pH 6 it readily self-assembles into long fibers. At higher concentrations the fibers form self-healing physical gels. Optimal gel strength and self-healing are found at a pH of around 7. The modulus of a 2 wt % gel at pH 7 is G' = 1700 Pa. Being protein-based, and amenable to further sequence engineering, we expect that these proteins are promising scaffold materials to be developed for a broad range of biomedical applications.
Understanding of the self- and co-assembly behavior of recombinant protein polymers : from design to implementation
Golinska, M.D. - \ 2014
Wageningen University. Promotor(en): Martien Cohen Stuart, co-promotor(en): Renko de Vries; Frits de Wolf. - Wageningen : Wageningen University - ISBN 9789461738134 - 165
biopolymeren - eiwitten - zelf-assemblage - recombinant eiwitten - elastine - collageen - enzymatische cross-linking - glutaminase - biopolymers - proteins - self assembly - recombinant proteins - elastin - collagen - enzymatic cross-linking - glutaminase

A key part of the growing field of biomedical sciences deals with the development of new, controlled and biocompatible biomaterials. In this thesis we present results on the design, production, purification and characterization of stimuli responsive protein polymers that could ultimately be used in that field. Protein-polymers are composed of two or three blocks that are able to self- and co-assemble. An important theme in the thesis is to highlight the specific advantages of our new protein polymer for future biomedical applications.

We have used recombinant DNA techniques and expression in methylotrophic yeast Pichia pastorisfor protein production. DNAs encoding various polypeptide blocks were designed, produced using general molecular biology techniques and combined into synthetic genes for protein polymers. Synthetic genes were cloned into P. pastorisexpression vector pPIC9 that integrates into the yeast genome. Yields were high, typically reaching gram-per-liter (of medium).

In Chapter 2we study dual-stimuli (pH, temperature) responsive silk-elastin-like protein polymers (SELPs). These polymers were designed to self- and co-assembly, controlled by both pH and temperature. The first protein is a diblock S24E40composed of 24 silk-like (S) repeats and 40 elastin-like (E) repeats. The other protein is a triblock S12C4E40, in which the Sand Eblocks are separated by a random coil block (C4) that serves as an inert ‘spacer’.

A C2SHSHC2protein polymer, which consists of a pH responsive, positively charged silk-like middle block SH, flanked by two random coil collagen-like blocks C2was studied in Chapter 3.

For this protein have studied fibril formation and gelling properties at pH values close to neutral, that are crucial for biomedical applications. We find that at physiological pH, these proteins form self-healing physical gels that fulfill many requirements for use in biomedical applications.

In Chapter 4we test the influence of enzymatic cross-linking on elasticity andmechanical properties ofhydrogels that include collagen-like domains, using microbial transglutaminase (mTGase) as an enzymatic crosslinker that catalyzes the coupling of glutamines to lysines. We show that even though the collagen-like blocks are not particularly good substrates for the mTGase, the few cross-links that are made have a strong effect on the physical properties of the protein-polymer hydrogels. For silk-collagen fiber gels, the elastic moduli can be increased by a factor of five, and for thermosensitive collagen hydrogels, the enzymatic cross-linking induces qualitatively new behavior, namely shape-memory of hydrogels.

Finally, we study the co-assembly of very asymmetric diblock copolymers with oppositely charged sodium poly(acrylic acid) (NaPAA) with a range of molar masses (Chapter 5). This asymmetric diblock consists of a cationic block of 12 lysines connected to a long (400 amino acid) collagen-like block with a net charge that is nearly zero. For shorter Na-PAA chains, spherical complex coacervates micelles are formed, as have been studied before in our lab. But, for long Na-PAA chains a new self-assembled structure is found: a single (Na-PAA) chain pearl-necklace of complex-coacervate micelles.

The general discussion of the thesis in Chapter 6, focuses on recombinant and natural hydrogels as biomaterials. We point out the specific advantages of recombinant proteins and also indicate where these still need to be improved in order to be used in biomedical applications. Finally, we make some suggestions for further research in this area.

Pearl-necklace complexes of flexible polyanions with neutral-cationic diblock copolymers
Golinska, M.D. ; Wolf, F.A. de; Cohen Stuart, M.A. ; Hernandez Garcia, A. ; Vries, R.J. de - \ 2013
Soft Matter 9 (2013)28. - ISSN 1744-683X - p. 6406 - 6411.
polyelectrolyte complexes - core micelles - poor solvent - adsorption - stability - brushes - films
We study the complexation of very asymmetric diblock copolymers (consisting of a cationic block of 12 lysines connected to a 400 amino acid long hydrophilic polypeptide block with a net charge that is nearly zero) with oppositely charged sodium poly(acrylic acid) (NaPAA) with a range of molar masses between 2 and 1300 kg mol-1. For shorter Na-PAA chains, spherical complex coacervate micelles are formed, but for long Na-PAA chains, with molar masses in excess of 250 kg mol-1, atomic force microscopy indicates the presence of pearl-necklace structures. Complexes most likely consist of only a single NaPAA chain, complexed to multiple diblocks. Hence, the size of the complexes can be fully controlled via the NaPAA molar mass. The occurrence of pearl-necklace complexes at higher NaPAA molar masses is attributed to the packing frustration that arises due to the small size of the cationic block of the diblock copolymers
Fibril Formation by pH and Temperature Responsive Silk-Elastin Block Copolymers
Golinska, M.D. ; Pham, T.T.H. ; Werten, M.W.T. ; Wolf, F.A. de; Cohen Stuart, M.A. ; Gucht, J. van der - \ 2013
Biomacromolecules 14 (2013)1. - ISSN 1525-7797 - p. 48 - 55.
protein-based polymers - periodic polypeptides - biosynthesis - biomaterials - purification - hydrogels - delivery - sequence - behavior - fusion
In this report, we study the self-assembly of two silk-elastin-like proteins: one is a diblock S24E40 composed of 24 silk-like (S) repeats and 40 elastin-like (E) repeats; the other is a triblock S12C4E40, in which the S and E blocks are separated by a random coil block (C4). Upon lowering the pH, the acidic silk-like blocks fold and self-assemble into fibrils by a nucleation-and-growth process. While silk-like polymers without elastin-like blocks form fibrils by heterogeneous nucleation, leading to monodisperse populations, the elastin-like blocks allow for homogeneous nucleation, which gives rise to polydisperse length distributions, as well as a concentration-dependent fibril length. Moreover, the elastin-like blocks introduce temperature sensitivity: at high temperature, the fibrils become sticky and tend to bundle and aggregate in an irreversible manner. Concentrated solutions of S12C4E40 form weak gels at low pH that irreversibly lose elasticity in temperature cycling; this is also attributed to fibril aggregation.
Tuning of Collagen Triple-Helix Stability in Recombinant Telechelic Polymers
Silva, C.I. da; Skrzeszewska, P.J. ; Golinska, M.D. ; Werten, M.W.T. ; Eggink, G. ; Wolf, F.A. de - \ 2012
Biomacromolecules 13 (2012)5. - ISSN 1525-7797 - p. 1250 - 1258.
triblock copolymers - stabilization - hydrogels - gelatin - gels - hydroxylation - proteins - kinetics - network - water
The melting properties of various triblock copolymers with random coil middle blocks (100–800 amino acids) and triple helix-forming (Pro-Gly-Pro)n end blocks (n = 6–16) were compared. These gelatin-like molecules were produced as secreted proteins by recombinant yeast. The investigated series shows that the melting temperature (Tm) can be genetically engineered to specific values within a very wide range by varying the length of the end block. Elongation of the end blocks also increased the stability of the helices under mechanical stress. The length-dependent melting free energy and Tm of the (Pro-Gly-Pro)n helix appear to be comparable for these telechelic polymers and for free (Pro-Gly-Pro)n peptides. Accordingly, the Tm of the polymers appeared to be tunable independently of the nature of the investigated non-cross-linking middle blocks. The flexibility of design and the amounts in which these nonanimal biopolymers can be produced (g/L range) create many possibilities for eventual medical application.
Microbiota composition of commercial cocoa powder from different geographical origins
Líma, L.J.R. ; Golinska, M.D. ; Nout, M.J.R. ; Zwietering, M.H. - \ 2008
In: Programme and Abstract Book, The 21st International ICFMH Symposium, ¿Evolving Microbial Food Quality and Safety¿, 1 - 4 September, 2008, Aberdeen, Scotland. - Aberdeen : IUMS - ICFMH - p. 177 - 177.
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