Theses and Dissertations

Date of Award

5-2022

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biochemistry and Molecular Biology

First Advisor

Dr. HyeongJun Kim

Second Advisor

Dr. Ahmed Touhami

Third Advisor

Dr. Michael Persans

Abstract

Faithful chromosome segregation involves ParB, a DNA-binding and compacting protein that specifically recognizes parS DNA sites near the replication origin. ParB spreads 10-20 kb from parS sites, which recruitments additional ParB molecules to neighboring DNA, forming higher-order nucleoprotein complexes. ParB is also a novel CTPase enzyme and hydrolyses CTP to self-dimerization and create a clamp that slides along DNA. To understand the roles of CTP in Bacillus subtilis ParB (BsParB), we purified wild type ParB (BsParB (WT)) and recombinant ParB (BsParB(R80A)) mutant. BsParB(R80A) was known to prevent CTP binding and not compact DNA. Proteins N- and C-terminally appended with a lysine-cysteine-lysine (KCK) tag were tested. We performed single-molecule DNA flow-stretching experiments both in the absence and presence of CTP. Both CTP binding and CTP hydrolysis impacts ParB-mediated DNA compaction, with parS sites modulating compaction rates, while BsParB(R80A) could compact DNA in a CTP-dependent manner. The KCK tag influenced DNA compaction.

Comments

Copyright 2022 Miranda Molina. All Rights Reserved.

https://go.openathens.net/redirector/utrgv.edu?url=https://www.proquest.com/dissertations-theses/exploring-dna-compaction-via-bacillus-subtilis/docview/2697658426/se-2?accountid=7119

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