Engineering Conferences International

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Ghanem, Fred

Tuesday, May 24 2022                                                     Session 3:   New Technologies for Enzyme Engineering 
IMMOBILIZED ENZYMES FOR GREEN PHARMACEUTICAL APPLICATIONS 
 
Fred Ghanem, Purolite, USA 
fred.ghanem@purolite.com 
 
 
With industry moving towards greener processes, biocatalysis is gaining more importance and more enzyme 
sequences are tailored every day for specific processes.  
 
Advantages of enzyme immobilization, parameters affecting the performance of immobilized enzymes and the 
types of processes employing such biocatalysts are being presented.  
 
The essential characteristics of synthetic enzyme carriers specifically designed to immobilize enzymes by 
affinity, covalent and hydrophobic interaction and their impact on the process performance are being discussed.  
 
To conclude, real-world case studies highlighting both novel and established industrial applications of 
immobilized enzymes used in the pharmaceutical industry are being presented. 
 
 
 
 


English

Immobilized enzymes for green pharmaceutical applications

Ellington, Andrew

Sunday, May 22 2022                                                                                                                            Plenary Talk 
USING MACHINE LEARNING TO IMPROVE PROTEIN FUNCTION 
 
Andrew D. Ellington, Raghav Shroff, Danny Diaz, James Loy,  
Austin Cole, Anastasia Kulikova, Tianlong Chen, Claus Wilke  
The University of Texas at Austin, USA 
ellingtonlab@gmail.com 
 
 
Machine learning methods have had a huge aspect on many aspects of our existence, and most recently have 
made great inroads into biotechnology.  To truly take advantage of these methods, very large data sets (for so-
called ‘deep learning’) are frequently required.  It can be daunting as biotechnologists to attempt to either 
develop experimental datasets for machine learning, or to promulgate methods that can be accurately and 
usefully applied to biological engineering.  To this end and building off the successes of Torng and Altman 
(2017), we have trained self-supervised 3D convolutional neural networks (3DCNN) and now 3D residual neural 
networks (3DResNets), that can leverage the enormous amount of data present in the Protein DataBank (PDB) 
to re-predict individual amino acids in protein structures with accuracies of up to 70%-85%.  We have also 
recently trained dense neural networks that combine predictions from structure-based neural networks with 
sequence-based neural networks (Transformers) and can speak to the relative performances separately and 
together.  What is especially interesting, though, is not re-prediction of wild-type residues, but the opportunity to 
leverage deep learning’s ability to find patterns in evolution (Transformers) and/or protein chemistry (3DCNNs) 
to identify positions where mutations may be beneficial.  In this regard, we will demonstrate how 3D CNNs have 
been used to functionally improve a variety of biotechnologically important proteins, including polymerases and 
esterases. 
 


Using machine learning to improve protein function

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Neuman, Thomas

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-19-2022 
Industry perspective on the future of organ-on-chip applications 
Thomas Neuman 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
 Thursday, May 19 2022                                                                                                                              Keynote 
INDUSTRY PERSPECTIVE ON THE FUTURE OF ORGAN-ON-CHIP APPLICATIONS 
 
Thomas Neuman, Nortis, USA 
Neumann@nortisbio.com 


Industry perspective on the future of organ-on-chip applications

Cicoira, Fabio

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-19-2022 
Materials and devices for stretchable and self-healing 
bioelectronics 
Fabio Cicoira 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
MATERIALS AND DEVICES FOR STRETCHABLE AND SELF-HEALING BIOELECTRONICS 
 
Fabio Cicoira, Polytechnique Montréal, Canada 
fabio.cicoira@polymtl.ca 
 


Materials and devices for stretchable and self-healing bioelectronics

Hwang, Jeonming

Kiick, Kristi L.

Sullivan, Millicent O.

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-19-2022 
Collagen-mimetic peptides for delivery of therapeutics in chronic 
wounds healing application 
Jeonming Hwang 
Kristi L. Kiick 
Millicent O. Sullivan 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
COLLAGEN-MIMETIC PEPTIDES FOR DELIVERY OF THERAPEUTICS IN CHRONIC WOUNDS HEALING 
APPLICATION 
 
Jeongmin Hwang, Department of Biomedical Engineering, University of Delaware, USA 
jmhwang@udel.edu 
Kristi L. Kiick, Department of Materials Science and Engineering, University of Delaware, USA 
Millicent O. Sullivan, Department of Chemical and Biomolecular Engineering, University of Delaware, USA 
 
 
Key Words: Collagen mimetic peptides, Gene/Drug delivery, Growth factor, Wound infection, Chronic wounds 
 
The enormous economic and societal burdens associated with chronic wounds suggest a critical need for new 
approaches to manipulate the wound microenvironment to support improved repair and prevent secondary 
complications. The main factors that lead to chronic wounds are the lack of growth factors to stimulate cellular 
healing responses, as well as the concomitant occurrence of wound infections that cause prolonged 
inflammation. Current treatments include wound dressings, topical application of growth factors or antibiotics, 
and various combination therapies that increase the accessibility of growth factors while eradicating 
microorganisms in the wound bed. However, the harsh wound environment often clears topically administered 
therapeutics from the wound, which reduces the local concentration of therapeutics and thus their effectiveness. 
The overall objective of our work is to improve the efficacy of topically administered therapeutics using peptide-
mediated interactions between nanocarriers and matrices. We specifically utilize collagen mimetic peptides 
(CMP) to tether nanostructure carriers onto collagen-containing matrices through the strand invasion of CMPs 
into native collagen, resulting in control over the delivery of the cargo in response to cell-mediated collagen 
degradation.  
 
Our studies have revealed that in general, nanocarriers modified with CMPs enable extended control of cargo 
release. Nanoparticles comprising EMP-CMP (Elastin-Mimetic Peptide and Collagen-Mimetic Peptide) 
conjugates facilitated the controlled, zero-order release of the antibiotic vancomycin, compared to the first-order 
release of vancomycin by a commonly used, liposomal drug nanocarrier. Furthermore, nanocarriers with CMP 
modification could be sequestered in collagen containing matrices to a greater extent than nanocarriers without 
CMP modification. As a result, the CMP-modified nanocarriers showed sustained release profiles. Also, CMP 
modification of polyethyleneimine (PEI)/DNA nanocomplexes, when sequestered within collagen-containing 
matrices, enhanced overall gene expression for at least 7 days. 
 
Additionally, our studies have demonstrated that the controlled delivery of therapeutics driven by CMP 
modification can also control their spatiotemporal action in vitro. The delivery of vascular endothelial growth 
factor (VEGF)-A via the use of CMP-modified PEI/DNA in collagen-containing matrices enhanced cellular 
signaling; murine fibroblasts differentiated to pro-healing myofibroblasts for at least 7 days and murine 
endothelial cells formed complete tubular networks with significantly greater volume and larger diameter than 
matrices lacking the CMP modification. Moreover, the delivery of vancomycin using EMP-CMP-nanoparticles 
tethered in collagen-containing matrices improved the half-life of vancomycin activity, resulting in the prevention 
of growth of methicillin-resistant Staphylococcus aureus (MRSA). Therefore, CMP modification of nanocarriers 
enables not only the controlled release of therapeutics from collagen-containing matrices through CMP retention 
within collagen, but also modulation of the bioactivities of therapeutics in vitro. Future work will include in vivo 
excisional wound studies to evaluate the efficacy of therapeutics, delivered by CMP-tethered nanocarriers, on 
wound healing, and evaluation of the platform for tissue regeneration applications.   
 
 
Figure - Schematic representation of 
collagen-mimetic peptide (CMP)-modified 
nanocarriers tethered to collagen-based 
matrices (Created with Biorender.com). 
 


Collagen-mimetic peptides for delivery of therapeutics in chronic wounds healing application

Lim, Roderick

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-17-2022 
Diffusion, disorder and dynamics in the nuclear pore complex 
Roderick Lim 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
DIFFUSION, DISORDER AND DYNAMICS IN THE NUCLEAR PORE COMPLEX 
 
Roderick Lim, Biozentrum, University of Basel, Switzerland 
roderick.lim@unibas.ch 
 
 


Diffusion, disorder and dynamics in the nuclear pore complex

Zhao, Yimu

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-16-2022 
Changes in extracellular matrix in failing human non-ischemic and 
ischemic hearts with mechanical unloading 
Yimu Zhao 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
CHANGES IN EXTRACELLULAR MATRIX IN FAILING HUMAN NON-ISCHEMIC AND 
ISCHEMIC HEARTS WITH MECHANICAL UNLOADING 
 
Yimu Zhao, University of Toronto, Canada 
elisa.cimetta@unipd.it 


Changes in extracellular matrix in failing human non-ischemic and ischemic hearts with mechanical unloading

Cimetta, Elisa

Engineering Conferences International 
ECI Digital Archives 
Nanotechnology In Medicine III: Enabling Next 
Generation Therapies Proceedings 
5-16-2022 
Microscale technologies to decode EV-mediated cell behavior 
Elisa Cimetta 
Follow this and additional works at: https://dc.engconfintl.org/nanotech_med_iii 
MICROSCALE TECHNOLOGIES TO DECODE EV-MEDIATED CELL BEHAVIOR 
 
Elisa Cimetta, University of Padova, Italy 
elisa.cimetta@unipd.it 


Microscale technologies to decode EV-mediated cell behavior

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Mohamed Ebrahim Alhamer Almulla, Fatma

Keshavarz-Moore, Eli

Engineering Conferences International 
ECI Digital Archives 
Microbial Engineering II Proceedings 
4-3-2022 
Optimisation of an alkaline lysis process for a plug- and play 
plasmid DNA production 
Fatma Mohamed Ebrahim Alhamer Almulla 
Eli Keshavarz-Moore 
Follow this and additional works at: https://dc.engconfintl.org/microbial_ii 
OPTIMISATION OF AN ALKALINE LYSIS PROCESS FOR A PLUG- AND PLAY PLASMID DNA 
PRODUCTION 
 
Fatma Mohamed Ebrahim Alhamer Almulla, University College London, United Kingdom 
Fatma.almulla.17@ucl.ac.uk 
Eli Keshavarz-Moore, University College London, United Kingdom 
 
 
Renewed interest in plasmid DNA (pDNA) production is the result of its role in the supply chain of novel 
biopharmaceuticals.” pDNA lies at the heart of viral and mRNA vector production because it provides the coding 
sequences for gene-based advanced therapy medicinal products (ATMPs). Its manufacture is therefore critical 
to both the supply and success of these products”1.  
 
One main bottleneck in processing of pDNA is the lysis step that still rely on techniques developed for 
laboratory-based DNA extraction. A robust process adaptable to a range of DNA sizes and scales remain to be 
fully realised. These include control of the alkaline environment, potential heterogeneity in a large vessel, 
change in the rheological properties of the process fluid and potentially shear impact if the molecule is large. 
Here, we focused on a batch lysis operation in a 1 L reactor. Three parameters studied were pH, mixing time 
and agitation speed. Using a central composite design for optimisation of these factors, we obtained the 
maximum plasmid yield (13 mg/gcells, R2=92%) and supercoiling content (91%, R2=95%) at 0.2M NaOH, 
mixing time of 15 minutes and tip speed of 2.3 m/s. These results will benchmark the design of plug- and- play 
platforms. 
 
 
1.T. Hitchcock 2021, Genetic Engineering & Biotechnology News 2nd August. 
 
 


Optimisation of an alkaline lysis process for a plug- and play plasmid DNA production

Fohler, Lisa

Engineering Conferences International 
ECI Digital Archives 
Microbial Engineering II Proceedings 
4-3-2022 
mDOE for development of optimized fed-batch cultivation for the 
production of PET degrading enzymes in E. coli 
Lisa Fohler 
Follow this and additional works at: https://dc.engconfintl.org/microbial_ii 
mDOE FOR DEVELOPMENT OF OPTIMIZED FED-BATCH CULTIVATION FOR THE PRODUCTION OF PET 
DEGRADING ENZYMES IN E. COLI 
 
Lisa Fohler, Austrian Center of Industrial Biotechnology (ACIB), University of Natural Resources and Life 
Sciences (BOKU), Austria 
lisa.fohler@students.boku.ac.at 
 
 
Key Words: enzymes, PET degradation, sustainability, mDOE, recycling 
 
Plastic production increases exponentially, with an estimated 448 million tonnes manufactured in 2015, about 
200 times more than in 1950. About 80 % of all produced plastic accumulates in landfills eventually, and natural 
degradation takes hundreds of years [1]. PET, the largest plastic waste fraction, is typically recycled by thermo-
mechanical means, resulting in a loss of mechanical stability. A more attractive alternative for recycling of PET 
waste is the enzymatically catalysed depolymerization und subsequent production of virgin PET [2]. The key to 
this recycling approach is the availability of sufficient amounts of thermostable, PET-degrading enzymes, which 
implies extremely high yield and cost efficiency requirements on the enzyme production processes used. So far, 
the production of PETases proofed difficult, with literature describing expression conditions of 16 °C to prevent 
rapid cell lysis due to assumed cell membrane degradation by the enzyme [3]. Expression temperatures this low 
are not feasible for big-scale production, therefore a new and more viable process has to be established. To 
tackle this challenge we identified three key tasks: cloning and selection of an expression system with which 
high titres can be achieved, set-up and optimization of fed-batch production using mDOE and lastly, 
implementing the gathered knowledge of the previous task to produce the enzyme in continuous culture. For the 
first task, the selected enzyme, PHL7, was cloned in the enGenes pENGX plasmid, equipped with N-terminal 
signal sequence for periplasmic expression and expression enhance tag and C-terminal 6His-tag for purification 
and analytics. The performance of 5 different signal sequences (spa, ompA, lamB, pelB and dsbA) was 
evaluated and the best performing leader selected. In addition, the influence of the C-terminal 6His-tag on 
expression and enzyme activity was examined. The expression host, E. coli enGenes X-press V2, was selected 
for its growth decoupling properties, redirecting all resources of the cell to recombinant protein production upon 
induction [4]. For the second task, set-up and optimization of fed-batch production, the critical input parameters 
and their corner points for the mDOE were identified in µ-scale bioreactor cultivation. These parameters were 
then used for a machine-learning aided process optimization based on a 33 fractional factorial design to 
maximize product titre of active enzyme. The insight about the expression behaviour will be used to implement a 
stable continuous production process, since we see it as a promising way to produce sufficient amounts of PET 
degrading enzymes. 
 
 
 
 
References: 
[1] R. Geyer, J. R. Jambeck, and K. L. Law, “Production, use, and fate of all plastics ever made,” Sci. Adv., 
vol. 3, no. 7, pp. 19–24, 2017. 
 
[2] V. Tournier et al., “An engineered PET depolymerase to break down and recycle plastic bottles,” Nature, 
vol. 580, no. 7802, pp. 216–219, 2020, doi: 10.1038/s41586-020-2149-4. 
 
[3] C. Sonnendecker et al., “Low Carbon Footprint Recycling of Post‐Consumer PET Plastic with a 
Metagenomic Polyester Hydrolase,” ChemSusChem, 2022, doi: 10.1002/cssc.202101062. 
 
[4] P. Stargardt, L. Feuchtenhofer, M. Cserjan-Puschmann, G. Striedner, and J. Mairhofer, “Bacteriophage 
Inspired Growth-Decoupled Recombinant Protein Production in Escherichia coli,” ACS Synth. Biol., vol. 9, no. 6, 
pp. 1336–1348, 2020, doi: 10.1021/acssynbio.0c00028. 
 


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