Wei‐Shou Hu

University of Minnesota

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Catherine M. Verfaillie

KU Leuven

Frank B. Cerra

University of Minnesota

Madhusudan V. Peshwa

MaxCyte (United States)

David H. Sherman

University of Michigan–Ann Arbor

Li‐Hong Malmberg

AbbVie (United States)

Pródromos Daoutidis

University of Minnesota

Scott L. Nyberg

Mayo Clinic in Arizona

Kelly K. Frame

University of Minnesota

Kartik Subramanian

Center for Systems Biology

Rory P. Remmel

University of Minnesota

Cooperative Institutions

University of Minnesota

219

John Wiley & Sons (United States)

111

Hudson Institute

Bioprocessing Technology Institute

Twin Cities Orthopedics

KU Leuven

Agency for Science, Technology and Research

National University of Singapore

University of Minnesota System

University of Minnesota Medical Center

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Metabolic Engineering of Cephalosporin Biosynthesis in Streptomyces clavuligerusa

Annals of the New York Academy of Sciences (1996)

Anurag Khetan Li‐Hong Malmberg David H. Sherman Wei‐Shou Hu

The biosynthesis of beta-lactams is one of the most thoroughly studied antibiotic pathways. The availability of the characteristics and the time profiles of activities of enzymes involved in the biosynthesis allows one to critically evaluate the potential rate-limiting steps in its production. Our approach to understanding the control of beta-lactam biosynthesis has been pursued using a two-stage strategy: (1) to predict the rate-limiting steps using a kinetic model and (2) to relax the rate-limiting steps by engineering the biosynthetic pathway or by altering the kinetic parameters of the predicted key rate-limiting enzyme. Kinetic analysis of the pathway dynamics of cephamycin C production in Streptomyces clavuligerus was performed using data obtained from wild type. Sensitivity analysis revealed that the availability of precursor alpha-aminoadipic acid and activity of ACV synthetase were the potential rate-limiting steps. Relaxation of the precursor limitation was accomplished by integration of an additional copy of the gene encoding lysine-epsilon-aminotransferase (lat) into the chromosome. The recombinant strain showed an increased level of cephamycin C production as expected. The intracellular levels of different intermediates in the pathway in batch cultures were analyzed.

Chemical technology

10.1111/j.1749-6632.1996.tb40543.x

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Citations (10)

Epigenomic features revealed by ATAC‐seq impact transgene expression in CHO cells

Biotechnology and Bioengineering (2021)

Zion Lee Marina Raabe Wei‐Shou Hu

Different regions of a mammalian genome have different accessibilities to transcriptional machinery. The integration site of a transgene affects how actively it is transcribed. Highly accessible genomic regions called super-enhancers have been recently described as strong regulatory elements that shape cell identity. Super-enhancers have been identified in Chinese hamster ovary (CHO) cells using the Assay for Transposase-Accessible Chromatin Sequencing (ATAC-seq). Genes near super-enhancer regions had high transcript levels and were enriched for oncogenic signaling and proliferation functions, consistent with an immortalized phenotype. Inaccessible regions in the genome with low ATAC signal also had low transcriptional activity. Genes in inaccessible regions were enriched for remote tissue functions such as taste, smell, and neuronal activation. A lentiviral reporter integration assay showed integration into super-enhancer regions conferred higher reporter expression than insertion into inaccessible regions. Targeted integration of an IgG vector into the Plec super-enhancer region yielded clones that expressed the immunoglobulin light chain gene mostly in the top 20% of all transcripts with the majority in the top 5%. The results suggest the epigenomic landscape of CHO cells can guide the selection of integration sites in the development of cell lines for therapeutic protein production.

Epigenomics

10.1002/bit.27701

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Determinants and rate laws of growth and death of hybridoma cells in continuous culture

Biotechnology and Bioengineering (1998)

An‐Ping Zeng Wolf‐Dieter Deckwer Wei‐Shou Hu

Experimental data from six hybridoma cell lines grown under diverse experimental conditions in both normal continuous and perfusion cultures are analyzed with respect to the significance of nutrients and products in determining the growth and death rates of cells and with respect to their mathematical modeling. It is shown that neither nutrients (glucose and glutamine) nor the common products lactic acid, ammonia, and monoclonal antibody can be generally assumed to be the clear-limiting or inhibiting factors for most of the cultures. Correspondingly, none of the unstructured models existing in the literature can be generally applied to describe the experimental data obtained over a relatively wide range of cultivation conditions as considered in this work. Surprisingly, for all cultures the specific growth rate (μ) almost linearly correlates with the ratio of the viable cell concentration (NV) to the dilution (perfusion) rate (D). Similarly, the specific death rate (kd) is a function of the ratio of the total cell concentration (Nt) to the dilution (perfusion) rate. These results strongly suggest the formation of not yet identified critical factors or autoinhibitors that determine both the growth and death rates of hybridoma cells. Based on these observations, simple kinetic models are developed for μ and kd which describe the experimental data satisfactorily. Analysis of the experimental data with the kinetic models reveals that under the current cultivation conditions the formation rate of the autoinhibitor(s) or the sensitivity of cell growth and death to the autoinhibitor(s) is mainly affected by the medium composition. Irrespective of the cell lines, cells grown on serum-containing media have almost the same model parameters, which are distinctively different from those of cells grown on serum-free media. Furthermore, in contrast to the prevailing view, kd is shown to positively correlate with μ if the effects of cell concentration and dilution (perfusion) rate are considered. Several important implications of these findings are discussed for the optimization and control of animal cell culture. © 1998 John Wiley & Sons, Inc. Biotechnol Bioeng 57: 642–654, 1998

Dilution

10.1002/(sici)1097-0290(19980320)57:6<642::aid-bit2>3.3.co;2-w

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Non-Linear Model Reduction for Metabolic Networks with Multiple Time-Scales

Ziomara P. Gerdtzen Pródromos Daoutidis Wei‐Shou Hu

We present a method for obtaining non-stiff nonlinear reduced-order models for metabolic networks, which exhibit dynamics in multiple time scales. The method is based on the successive application of singular perturbation arguments, starting from the fastest time scale and proceeding to the slowest one. The method is successfully applied to a detailed model of central carbon metabolism in human erythrocytes

Nonlinear dynamical systems

10.1109/.2005.1467069

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Transcriptome and proteome analysis of Chinese hamster ovary cells under low temperature and butyrate treatment

Journal of Biotechnology (2009)

Anne Kantardjieff Nitya M. Jacob Joon Chong Yee Eyal Epstein Yee Jiun Kok

10.1016/j.jbiotec.2009.09.008

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Citations (141)

Mechanisms of peptide sex pheromone regulation of conjugation in Enterococcus faecalis

MicrobiologyOpen (2017)

Yuqing Chen Arpan Bandyopadhyay Briana Kozlowicz Heather A. H. Haemig Albert Tai

Abstract In many gram positive bacteria, horizontal transfer and virulence are regulated by peptide‐mediated cell‐cell signaling. The heptapeptide cCF 10 ( C ) activates conjugative transfer of the Enterococcus faecalis plasmid pCF 10, whereas the iCF 10 ( I ) peptide inhibits transfer. Both peptides bind to the same domain of the master transcription regulator PrgX, a repressor of transcription of the prgQ operon encoding conjugation genes. We show that repression of prgQ by PrgX tetramers requires formation of a pCF 10 DNA loop where each of two PrgX DNA ‐binding sites is occupied by a dimer. I binding to PrgX enhances prgQ repression, while C binding has the opposite effect. Previous models suggested that differential effects of these two peptides on the PrgX oligomerization state accounted for their distinct functions. Our new results demonstrate that both peptides have similar, high‐binding affinity for PrgX, and that both peptides actually promote formation of PrgX tetramers with higher DNA ‐binding affinity than Apo‐PrgX. We propose that differences in repression ability of PrgX/peptide complexes result from subtle differences in the structures of DNA ‐bound PrgX/peptide complexes. Changes in the induction state of a donor cell likely results from replacement of one type of DNA ‐bound peptide/PrgX tetramer with the other.

Bacterial conjugation

Transcription

10.1002/mbo3.492

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Citations (27)

On‐line characterization of a hybridoma cell culture process

Biotechnology and Bioengineering (1994)

Weichang Zhou Wei‐Shou Hu

Abstract The on‐line determination of the physiological state of a cell culture process requires reliable on‐line measurements of various parameters and calculations of specific rates from these measurements. The cell concentration of a hybridoma culture was estimated on‐line by measuring optical density (OD) with a laser turbidity probe. The oxygen uptake rate (OUR) was determined by monitoring dynamically dissolved oxygen concentration profiles and closing oxygen balances in the culture. The base addition for neutralizing lactate produced by cells was also monitored on‐line via a balance. Using OD and OUR measurements, the specific growth and specific oxygen consumption rates were determined on‐line. By combining predetermined stoichiometric relationships among oxygen and glucose consumption and lactate production, the specific glucose consumption and lactate production rates were also calculated on‐line. Using these on‐line measurements and calculations, the hybridoma culture process was characterized on‐line by identifying the physiological states. They will also facilitate the implementation of nutrient feeding strategies for fed‐batch and perfusion cultures. © 1994 John Wiley & Sons, Inc.

Optical density

10.1002/bit.260440205

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Citations (111)

Kinetics of Cell Growth Processes

Wei‐Shou Hu

This chapter discusses the quantitative description of cell growth (and briefly cell death), as well as the mathematical description and the growth model that can be used to predict the kinetic behavior of cells in culture. As cells grow, they increase in number, mass, and size. In a natural environment or in culture medium, their growth rate can change in response to fluctuations in the concentration of nutrients, metabolites, oxygen, temperature, pH, and other factors. The energy needs required for cell growth are relatively constant, regardless of whether the yeast cell is at an oxidative metabolic state or a fermentative metabolic state. When growing cells, all of the essential nutrients must be provided. As in the case of cell death in culture, the process of killing cells by sterilization is typically considered to follow first-order kinetics with respect to live cells. The objective of sterilization is to eliminate live bacteria in the entire volume.

Sterilization

10.1002/9781119159056.ch5

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Cybernetic Modeling and Regulation of Metabolic Pathways in Multiple Steady States of Hybridoma Cells

Biotechnology Progress (2000)

M.J. Guardia Anshu Gambhir Anna Europa Doraiswami Ramkrishna Wei‐Shou Hu

Hybridoma cells utilize a pair of complementary and partially substitutable substrates, glucose and glutamine, for growth. It has been shown that cellular metabolism shifts under different culture conditions. When those cultures at different metabolic states are switched to a continuous mode, they reach different steady states under the same operating conditions. A cybernetic model was constructed to describe the complementary and partial substitutable nature of substrate utilization. The model successfully predicted the metabolic shift and multiple steady-state behavior. The results are consistent with the experimental observation that the history of the culture affects the resulting steady state.

Metabolic Regulation

Mode (computer interface)

10.1021/bp000069a

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Citations (28)

Effect of glutamate on the degradation of pentachlorophenol by Flavobacterium sp.

Applied Microbiology and Biotechnology (1991)

JorgeF. Gonzalez Wei‐Shou Hu

Pentachlorophenol

Degradation

10.1007/bf00180644

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Citations (37)