Title:Proteomic analysis of a hom-disrupted, cephamycin C overproducing Streptomyces clavuligerus
Volume: 28
Issue: 2
Author(s): Eser Ünsaldı, Aslıhan Kurt-Kızıldoğan, Servet Özcan, Dörte Becher, Birgit Voigt, Caner Aktaş and Gülay Özcengiz*
Affiliation:
- Department of Biological Sciences, Middle East Technical University, Ankara 06800,Turkey
Keywords:
Cephamycin C, Streptomyces clavuligerus, hom-disruption, proteomics, MALDI-TOF/MS, nanoLC-MS/MS.
Abstract:
Background: Streptomyces clavuligerus is prolific producer of cephamycin C, a medically
important antibiotic. In our former study, cephamycin C titer was 2-fold improved by disrupting
homoserine dehydrogenase (hom) gene of aspartate pahway in Streptomyces clavuligerus NRRL
3585.
Objective: In this article, we aimed to provide a comprehensive understanding at the proteome level
on potential complex metabolic changes as a consequence of hom disruption in Streptomyces
clavuligerus AK39.
Methods: A comparative proteomics study was carried out between the wild type and its hom disrupted
AK39 strain by 2 Dimensional Electrophoresis-Matrix Assisted Laser Desorption and Ionization
Time-Of-Flight Mass Spectrometry (2DE MALDI-TOF/MS) and Nanoscale Liquid Chromatography-
Tandem Mass Spectrometry (nanoLC-MS/MS) analyses. Clusters of Orthologous
Groups (COG) database was used to determine the functional categories of the proteins. The theoretical
pI and Mw values of the proteins were calculated using Expasy pI/Mw tool.
Results: “Hypothetical/Unknown” and “Secondary Metabolism” were the most prominent categories
of the differentially expressed proteins. Upto 8.7-fold increased level of the positive regulator
CcaR was a key finding since CcaR was shown to bind to cefF promoter thereby direcly controlling
its expression. Consistently, CeaS2, the first enzyme of CA biosynthetic pathway, was 3.3-
fold elevated. There were also many underrepresented proteins associated with the biosynthesis of
several Non-Ribosomal Peptide Synthases (NRPSs), clavams, hybrid NRPS/Polyketide synthases
(PKSs) and tunicamycin. The most conspicuously underrepresented protein of amino acid
metabolism was 4-Hydroxyphenylpyruvate dioxygenase (HppD) acting in tyrosine catabolism. The
levels of a Two Component System (TCS) response regulator containing a CheY-like receiver domain
and an HTH DNA-binding domain as well as DNA-binding protein HU were elevated while
a TetR-family transcriptional regulator was underexpressed.
Conclusion: The results obtained herein will aid in finding out new targets for further improvement
of cephamycin C production in Streptomyces clavuligerus.