TY  - JOUR
A1  - Altintas, Zeynep
A1  - Takiden, Aref
A1  - Utesch, Tillmann
A1  - Mroginski, Maria A.
A1  - Schmid, Bianca
A1  - Scheller, Frieder W.
A1  - Süssmuth, Roderich D.
T1  - Integrated approaches toward high-affinity artificial protein binders obtained via computationally simulated epitopes for protein recognition
JF  - Advanced functional materials
N2  - Widely used diagnostic tools make use of antibodies recognizing targeted molecules, but additional techniques are required in order to alleviate the disadvantages of antibodies. Herein, molecular dynamic calculations are performed for the design of high affinity artificial protein binding surfaces for the recognition of neuron specific enolase (NSE), a known cancer biomarker. Computational simulations are employed to identify particularly stabile secondary structure elements. These epitopes are used for the subsequent molecular imprinting, where surface imprinting approach is applied. The molecular imprints generated with the calculated epitopes of greater stability (Cys-Ep1) show better binding properties than those of lower stability (Cys-Ep5). The average binding strength of imprints created with stabile epitopes is found to be around twofold and fourfold higher for the NSE derived peptide and NSE protein, respectively. The recognition of NSE is investigated in a wide concentration range, where high sensitivity (limit of detection (LOD) = 0.5 ng mL(-1)) and affinity (dissociation constant (K-d) = 5.3 x 10(-11)m) are achieved using Cys-Ep1 imprints reflecting the stable structure of the template molecules. This integrated approach employing stability calculations for the identification of stabile epitopes is expected to have a major impact on the future development of high affinity protein capturing binders.
KW  - artificial protein binders
KW  - cancer markers
KW  - computationally simulated epitopes
KW  - molecular imprinting
KW  - protein recognition
Y1  - 2019
U6  - https://doi.org/10.1002/adfm.201807332
SN  - 1616-301X
SN  - 1616-3028
VL  - 29
IS  - 15
PB  - Wiley-VCH
CY  - Weinheim
ER  - 
TY  - JOUR
A1  - Reyna-González, Emmanuel
A1  - Schmid, Bianca
A1  - Petras, Daniel
A1  - Süssmuth, Roderich D.
A1  - Dittmann, Elke
T1  - Leader Peptide-Free In Vitro Reconstitution of Microviridin Biosynthesis Enables Design of Synthetic Protease-Targeted Libraries
JF  - Angewandte Chemie : a journal of the Gesellschaft Deutscher Chemiker ; International edition
N2  - Microviridins are a family of ribosomally synthesized and post-translationally modified peptides with a highly unusual architecture featuring non-canonical lactone as well as lactam rings. Individual variants specifically inhibit different types of serine proteases. Here we have established an efficient in vitro reconstitution approach based on two ATP-grasp ligases that were constitutively activated using covalently attached leader peptides and a GNAT-type N-acetyltransferase. The method facilitates the efficient in vitro one-pot transformation of microviridin core peptides to mature microviridins. The engineering potential of the chemo-enzymatic technology was demonstrated for two synthetic peptide libraries that were used to screen and optimize microviridin variants targeting the serine proteases trypsin and subtilisin. Successive analysis of intermediates revealed distinct structure-activity relationships for respective target proteases.
KW  - biosynthesis
KW  - cyanobacteria
KW  - microviridins
KW  - natural products
KW  - peptides
Y1  - 2016
U6  - https://doi.org/10.1002/anie.201604345
SN  - 1433-7851
SN  - 1521-3773
VL  - 55
SP  - 9398
EP  - 9401
PB  - Wiley-VCH
CY  - Weinheim
ER  - 
TY  - JOUR
A1  - Dehm, Daniel
A1  - Krumbholz, Julia
A1  - Baunach, Martin
A1  - Wiebach, Vincent
A1  - Hinrichs, Katrin
A1  - Guljamow, Arthur
A1  - Tabuchi, Takeshi
A1  - Jenke-Kodama, Holger
A1  - Süssmuth, Roderich D.
A1  - Dittmann-Thünemann, Elke
T1  - Unlocking the spatial control of secondary metabolism uncovers hidden natural product diversity in nostoc punctiforme
JF  - ACS chemical biology
N2  - Filamentous cyanobacteria belong to the most prolific producers of structurally unique and biologically active natural products, yet the majority of biosynthetic gene clusters predicted for these multicellular collectives are currently orphan. Here, we present a systems analysis of secondary metabolite gene expression in the model strain Nostoc punctiforme PCC73102 using RNA-seq and fluorescence reporter analysis. Our data demonstrate that the majority of the cryptic gene clusters are not silent but are expressed with regular or sporadic pattern. Cultivation of N. punctiforme using high-density fermentation overrules the spatial control and leads to a pronounced upregulation of more than 50% of biosynthetic gene clusters. Our data suggest that a combination of autocrine factors, a high CO2 level, and high light account for the upregulation of individual pathways. Our overarching study not only sheds light on the strategies of filamentous cyanobacteria to share the enormous metabolic burden connected with the production of specialized molecules but provides an avenue for the genome-based discovery of natural products in multicellular cyanobacteria as exemplified by the discovery of highly unusual variants of the tricyclic peptide microviridin.
Y1  - 2019
U6  - https://doi.org/10.1021/acschembio.9b00240
SN  - 1554-8929
SN  - 1554-8937
VL  - 14
IS  - 6
SP  - 1271
EP  - 1279
PB  - American Chemical Society
CY  - Washington
ER  -