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Screening for interaction partners of SepIVA and AfsK, proteins involved in polar growth of Streptomyces venezuelae

Konopka, Veronika (2021) MOBK01 20211
Degree Projects in Molecular Biology
Popular Abstract
Fishing for interaction partners

Bacteria, the most abundant living organisms, come in many shapes and sizes depending on how they grow and divide. Some bacteria like Escherichia coli form rodlike structures and divide in the middle. Others, like Streptomyces venezuelae, grow as long and branching filamentous cells called hyphae. They build their cell walls at the tips of these hyphae. Two proteins called SepIVA and AfsK are located at these hyphal tips and are believed to be involved in the growth of S. venezuelae. To understand the role of these proteins I aimed to identify interaction partners that potentially work together with SepIVA and AfsK.

I used a technique called the bacterial adenylate cyclase two-hybrid system. In this... (More)
Fishing for interaction partners

Bacteria, the most abundant living organisms, come in many shapes and sizes depending on how they grow and divide. Some bacteria like Escherichia coli form rodlike structures and divide in the middle. Others, like Streptomyces venezuelae, grow as long and branching filamentous cells called hyphae. They build their cell walls at the tips of these hyphae. Two proteins called SepIVA and AfsK are located at these hyphal tips and are believed to be involved in the growth of S. venezuelae. To understand the role of these proteins I aimed to identify interaction partners that potentially work together with SepIVA and AfsK.

I used a technique called the bacterial adenylate cyclase two-hybrid system. In this system, a gene coding for a protein called adenylate cyclase is split into two fragments. First, a bait was designed with the gene coding for the AfsK protein fused with another gene coding for a fragment of adenylate cyclase. Then the bait was used to fish for partners in a pool of genes each coding for a protein of S. venezuelae fused with the gene coding for the other fragment of adenylate cyclase. Active adenylate cyclase can turn on the production of different proteins, including those that allow the bacteria to use sugars lactose (Lac+) and maltose (Mal+) as an alternative to glucose energy source. When the bait protein interacts with another protein in the pool, the two fragments of adenylate cyclase are brought close together. The cells in which this occurs regain the adenylate cyclase activity and grow as blue colonies on a medium with lactose (Fig. 1A), and red colonies on a medium with maltose (Fig. 1B), as the only energy source.

In total, 12 bacterial colonies showed both Lac+ and Mal+ characteristics when fishing with the SepIVA bait. From these, eight potential interactions were identified, including one in which SepIVA interacted with itself. Among the putative interaction partners were three different transcription factors (proteins that control transcription from DNA to mRNA) and a kinase (a protein that allows the transfer of a phosphate molecule), to mention some. Further experiments are necessary to confirm and understand these interactions. Although no interactions were identified for AfsK, two positive baits were designed and can be used in future experiments to fish for interacting proteins. Besides laying the ground for future studies, these results provide an insight into the variety of potential interactions between the proteins involved in cell growth, thus presenting the complexity of this basic mechanism of life. Unraveling this complex machinery of cell growth in bacteria is valuable as this knowledge can for example be used in development of new antibiotics.

Supervisors: Klas Flärdh and Parminder Singh Mavi
Bachelor’s thesis project 15 credits in MOBK01, 2021
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Konopka, Veronika
supervisor
organization
course
MOBK01 20211
year
type
M2 - Bachelor Degree
subject
language
English
id
9059951
date added to LUP
2021-06-30 11:53:05
date last changed
2021-06-30 11:53:05
@misc{9059951,
  author       = {{Konopka, Veronika}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{Screening for interaction partners of SepIVA and AfsK, proteins involved in polar growth of Streptomyces venezuelae}},
  year         = {{2021}},
}