Cell Division

Arends, S J Ryan; Williams, Kyle B.; Kustusch, Ryan J.; Weiss, David S.

doi:10.1128/9781555815806.ch10

Cited by 2 publications

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“…Eventually, upon extended growth at the higher temperature, the cells would lyse and die. Now, some four decades later, cell division in E. coli is known to be mediated by over 20 proteins that localize to a ring like structure at the midcell, referred to as the septal ring or divisome (Figure 1.1, For recent reviews see: Goehring and Beckwith, 2005;Vicente and Rico, 2006;Arends et al, 2007). The septal ring is a large, complex, multi-protein structure that is responsible for many steps of the division process.…”

Section: Overview Of Cell Divisionmentioning

confidence: 99%

“…Some of the work presented in this thesis has been published (Arends et al, 2010). (Chen & Beckwith, 2001;Goehring & Beckwith, 2005) FtsI is a class B high molecular weight PBP (see Chapter 1 for more information on FtsI) that localizes to the division site, where it is essential for crosslinking newly synthesized PG at the septum (Adam et al, 1997;Weiss et al, 1999;Arends, 2007). In terms of structure, FtsI is a bitopic membrane protein with a small cytoplasmic domain, a single transmembrane (TM) helix, and a large periplasmic region that contains both a domain of unknown function and a transpeptidase catalytic domain (Bowler & Spratt, 1989;Goffin et al, 1996;Goffin & Ghuysen, 1998).…”

Section: An Overview Of This Thesismentioning

confidence: 99%

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The DamX cell division protein of Escherichia coli

Williams¹

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In the bacterium Escherichia coli, cell division involves the concerted inward growth of all three layers of the cell envelope: the cytoplasmic membrane, the peptidoglycan (PG) cell wall, and the outer membrane. This is a complex, highly regulated process that involves over 20 proteins. Four of these proteins contain a domain of ~70 amino acids known as a SPOR domain (Pfam no. 05036). One of these SPOR domains (from a protein named FtsN) has been shown previously to bind PG. In this thesis we show that six additional SPOR domains, three from E. coli and three from other bacterial species, also bind PG. Thus, PG binding is a general activity of SPOR domains.We then examine the SPOR domain from DamX of E. coli in detail. In collaboration with Dr. Andrew Fowler of the NMR Core Facility, we determined the solution structure of the domain. The domain adopts an "RNP fold," characterized by a four-stranded antiparallel β-sheet that is buttressed on one side by α-helixes. Several mutant forms of the DamX SPOR domain were constructed and studied both in vivo and in vitro. These studies support the following inferences: 1) The β-sheet is the PG-binding site; 2) The βsheet contains critical information for targeting the SPOR domain to the midcell; 3) The SPOR domain probably localizes to the midcell by binding preferentially to septal PG; and 4) It follows, then, that septal PG must differ from PG elsewhere around the cell. We suggest that further studies of the SPOR:PG interaction will yield novel insights into PG biogenesis during septation.This thesis also presents an in vivo characterization of several mutant forms of a cytoplasmic membrane protein named FtsW, homologs of which are found in all bacteria that contain a PG cell wall. FtsW recruits a PG synthase named FtsI to the division site and might also transport PG precursors across the cytoplasmic membrane. We systematically mutagenized each of FtsW's ten transmembrane (TM) helixes and investigated the ability of the mutant proteins to support division, localize to the division site, and recruit FtsI. This characterization leads us to propose that TM1 is involved in targeting FtsW to the division site, TM4 is involved in the putative transport activity, and TM10 is involved in recruitment of FtsI.Abstract Approved: _____________________________ Thesis Supervisor _____________________________

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Section: Overview Of Cell Divisionmentioning

confidence: 99%

Section: An Overview Of This Thesismentioning

confidence: 99%

The DamX cell division protein of Escherichia coli

Williams¹

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ATP-Binding Site Lesions in FtsE Impair Cell Division

2009

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FtsE and FtsX of Escherichia coli constitute an apparent ABC transporter that localizes to the septal ring. In the absence of FtsEX, cells divide poorly and several membrane proteins essential for cell division are largely absent from the septal ring, including FtsK, FtsQ, FtsI, and FtsN. These observations, together with the fact that ftsE and ftsX are cotranscribed with ftsY, which helps to target some proteins for insertion into the cytoplasmic membrane, suggested that FtsEX might contribute to insertion of division proteins into the membrane. Here we show that this hypothesis is probably wrong, because cells depleted of FtsEX had normal amounts of FtsK, FtsQ, FtsI, and FtsN in the membrane fraction. We also show that FtsX localizes to septal rings in cells that lack FtsE, arguing that FtsX targets the FtsEX complex to the ring. Nevertheless, both proteins had to be present to recruit further Fts proteins to the ring. Mutant FtsE proteins with lesions in the ATP-binding site supported septal ring assembly (when produced together with FtsX), but these rings constricted poorly. This finding implies that FtsEX uses ATP to facilitate constriction rather than assembly of the septal ring. Finally, topology analysis revealed that FtsX has only four transmembrane segments, none of which contains a charged amino acid. This structure is not what one would expect of a substrate-specific transmembrane channel, leading us to suggest that FtsEX is not really a transporter even though it probably has to hydrolyze ATP to support cell division.

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Cell Division

Cited by 2 publications

References 163 publications

The DamX cell division protein of Escherichia coli

The DamX cell division protein of Escherichia coli

ATP-Binding Site Lesions in FtsE Impair Cell Division

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