Proteomics Profiling of Chimeric-Truncated Tissue Plasminogen activator Producing- Chinese Hamster Ovary Cells Cultivated in a Chemically Defined Medium Supplemented with Protein Hydrolysates

Azarian, Bahareh; Sajedin, Seyedeh Matin; Azimi, Amin; Raigani, Mozhgan; Vaziri, Behrouz; Davami, Fatemeh

doi:10.18869/acadpub.ibj.21.3.154

Proteomics Profiling of Chimeric-Truncated Tissue Plasminogen activator Producing- Chinese Hamster Ovary Cells Cultivated in a Chemically Defined Medium Supplemented with Protein Hydrolysates

Authors

Bahareh Azarian ¹

Seyedeh Matin Sajedin ¹

Amin Azimi ¹

Mozhgan Raigani ²

Behrouz Vaziri ¹

Fatemeh Davami ¹

¹ Protein Chemistry Unit, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran

² Eukaryotic Expression Unit, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran

10.18869/acadpub.ibj.21.3.154

Abstract

Background: Culture media enrichment through the addition of protein hydrolysates is beneficial for achieving higher protein expression. Methods: In this study, designing the optimum mixture of four soy and casein-derived hydrolysates was successfully performed by design of experiment and specific productivity increased in all predicted combinations. Protein profile of recombinant CHO (rCHO) cells producing tissue plasminogen activator in a serum-free medium (SFM) supplemented with designed hydrolysate additives was compared to that of rCHO cells cultivated in SFM. Results: Identification of differentially expressed proteins using two-dimensional gel electrophoresis coupled with MALDI-TOF/TOF revealed the role of energy metabolism related proteins and importance of prevention of oxidative stress by this special media enrichment strategy. Up-regulation of mitochondrial enzymes, pyruvate dehydrogenase E1 and Peroxiredoxin-III, as well as other proteins involved in metabolic pathways, and uridine monophosphate/cytidine monophosphate kinase indicated higher metabolic activity. Furthermore, along with antioxidant effect of peptones, proteins with antioxidant function such as ferritin and peroxiredoxin-III were up-regulated. Conclusion: Understanding molecular mechanisms involved in enhancement of protein expression can provide new approaches for efficiently engineering rCHO cell. These results support the competence of proteomics studies in finding new insights to biochemical pathways for a knowledge-based optimization of media compositions.

Keywords

CHO Cells

DOE

Hydrolysates

MALDI-TOF/TOF

Proteomics