The Protein Journal

, Volume 37, Issue 2, pp 132–143 | Cite as

The Catalytically Inactive Mutation of the Ubiquitin-Conjugating Enzyme CDC34 Affects its Stability and Cell Proliferation

  • Xun Liu
  • Yang Zhang
  • Zhanhong Hu
  • Qian Li
  • Lu Yang
  • Guoqiang Xu


The ubiquitin proteasome system (UPS) plays important roles in the regulation of protein stability, localization, and activity. A myriad of studies have focused on the functions of ubiquitin ligases E3s and deubiquitinating enzymes DUBs due to their specificity in the recognition of downstream substrates. However, the roles of the most ubiquitin-conjugating enzymes E2s are not completely understood except that they transport the activated ubiquitin and form E2–E3 protein complexes. Ubiquitin-conjugating enzyme CDC34 can promote the degradation of downstream targets through the UPS whereas its non-catalytic functions are still elusive. Here, we find that mutation of the catalytically active cysteine to serine (C93S) results in the reduced ubiquitination, increased stability, and attenuated degradation rate of CDC34. Through semi-quantitative proteomics, we identify the CDC34-interacting proteins and discover that the wild-type and mutant proteins have many differentially interacted proteins. Detailed examination finds that some of them are involved in the regulation of gene expression, cell growth, and cell proliferation. Cell proliferation assay reveals that both the wild-type and C93S proteins affect the proliferation of a cancer cell line. Database analyses show that CDC34 mRNA is highly expressed in multiple cancers, which is correlated with the reduced patient survival rate. This work may help to elucidate the enzymatic and non-enzymatic functions of this protein and might provide additional insights for drug discovery targeting E2s.


Catalytic site CDC34 Cell proliferation Protein degradation Proteomics Ubiquitination 



Cell counting kit-8


Ubiquitin-conjugating enzyme E2 R1 or cell division cycle 34




Casein kinase I isoform γ2


Guanine nucleotide-binding protein-like 3


Human embryonic kidney


DNA replication licensing factor MCM7


Mass spectrometry


Negative control


Polymerase chain reaction




Peptide spectrum match




Sodium dodecyl sulfate–polyacrylamide gel electrophoresis


Standard error of measurements




Ubiquitin proteasome system


C93S mutant


CDC34 wild-type



We thank Yarong Wang at the Mass Spectrometry core facility of the Medical School of Soochow University for the assistance during the MS analysis. We also appreciate Dr. Xiaoyan Qiu at Soochow University for her critical reading of the manuscript. This work was supported by the National Natural Science Foundation of China (31670833 & 31700722), Postgraduate Research and Practice Innovation Program of Jiangsu Province (KYCX17_2040), Jiangsu Key Laboratory of Neuropsychiatric Diseases (BM2013003), a project funded by the Priority Academic Program Development (PAPD) of Jiangsu Higher Education Institutions.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

Human Participants and Animals

This article does not contain any studies with human participants or animals performed by any of the authors

Supplementary material

10930_2018_9766_MOESM1_ESM.docx (2.5 mb)
Supplementary material 1 (DOCX 2541 KB)
10930_2018_9766_MOESM2_ESM.xlsx (47 kb)
Supplementary material 2 (XLSX 46 KB)


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© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Jiangsu Key Laboratory of Neuropsychiatric Diseases and College of Pharmaceutical Sciences, Jiangsu Key Laboratory of Preventive and Translational Medicine for Geriatric DiseasesSoochow UniversitySuzhouChina

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