Faculty of Biological Sciences

Dr Christopher Thomas

BA, MA, Oxford; PhD, Leicester
Lecturer
School of Molecular and Cellular Biology

Background: Postdoctoral work at the Department of Biochemistry, University of Leicester; Wellcome Career Development Fellowship awarded 1994; moved to University of Leeds in 1995. Appointed Lecturer in 1998.

Contact:  Miall 6.02, +44(0) 113 34 33040, email address for  

You can read more about Dr Thomas's interests here:
www.astbury.leeds.ac.uk/People/staffpage.php?StaffID=CDT
www.fbs.leeds.ac.uk/staff/cdt/

Figure 1: The Rep proteins form specific homodimers prior to initiation of replication

Figure 2: Accessory protein MobC is required for MobA to cleave at oriT

Research Interests

Replication- and conjugation-specific proteins and the maintenance of extrachromosomal DNA

Bacterial resistance to antibiotics is often conferred by the presence of plasmids, or extrachromosomal DNA elements. Small plasmids of Gram positive organisms (such as Staphylococcus aureus) typically replicate via a 'rolling circle' mechanism, following cleavage of one DNA strand by a replication initiator protein. The mechanism of this process is related to that used by type-I topoisomerases and other replication initiators such as the gene II protein of bacteriophage M13.

The RepD protein initiates replication of plasmid pC221. We have characterised a new sequence motif at the active site of RepD, enabling it to carry out both DNA cleavage and religation reactions without added ATP. This motif does not, however, determine the dimerisation properties of the protein, which we are currently refining via domain swapping and site-directed mutagenesis experiments. RepD also recruits the PcrA helicase to the origin of replication, which serves to separate the DNA strands permitting replication. This interaction also stimulates the processivity of the helicase; we are investigating this mechanism using cloned RepD and PcrA studied in vitro using both biochemical and structural techniques.

Mobilisation of pC221 has been observed in the presence of self-transmissible plasmids. In addition to the relaxase protein MobA we have identified a crucial accessory protein, MobC, which is common to all plasmids of this family. Our current studies of the relaxase complex are directed towards understanding the roles of MobA and MobC in the specific recognition of the cognate transfer origin, oriT, and how they bring about the cleavage process.

 

Faculty Research and Innovation



Studentship information

Undergraduate project topics:

  • Typical laboratory projects on offer are concerned with MRSA and DNA replication; more recently students have also been working on plasmid replication in thermophilic bacteria. Projects are usually chosen by students looking to gain experience in molecular biology techniques (PCR, cloning etc.) but the precise content can usually be adjusted to suit the student's interests.

Postgraduate studentship areas:

  • Plasmid replication and transfer; maintenance of chromosomal DNA; protein:DNA and protein:protein interactions; development of novel targets for antibiotics.

See also:

Modules managed

BIOC1302 - Introductory Biochemistry: Practical Skills

Modules taught

BIOC1302 - Introductory Biochemistry: Practical Skills
BIOC1303 - Introductory Biochemistry: Problem Solving and Data Handling
BIOC2303 - Intermediate Biochemistry: Skills
BIOC3231/32/BIOL3211/MICR3212 c - ATU - DNA maintenance in pathogens
BIOL2301/03/MICR2320/BIOL3400 - BIOL/BIOC/MICR advanced skills/Skills in the Cell Biology of Human Disease
BIOL5294M - MSc Bioscience Research Project Proposal
BIOL5392M - Bioscience MSc Research Project
BMSC1213 - Basic Laboratory and Scientific Skills 2
MICR3080 - Microbiology Research Project
SPSC2203 - Exercise Biochemistry

Centre memberships:

Group Leader Dr Christopher Thomas  (Lecturer)

Replication- and conjugation-specific proteins and the maintenance of extrachromosomal DNA 

Machon C, Lynch GP, Thomson NH, Scott DJ, Thomas CD, Soultanas P RepD-mediated recruitment of PcrA helicase at the Staphylococcus aureus pC221 plasmid replication origin, oriD NUCLEIC ACIDS RES 38 1874-1888, 2010
DOI:10.1093/nar/gkp1153

Slatter AF, Thomas CD, Webb MR PcrA Helicase Tightly Couples ATP Hydrolysis to Unwinding Double-Stranded DNA, Modulated by the Initiator Protein for Plasmid Replication, RepD BIOCHEMISTRY-US 48 6326-6334, 2009
DOI:10.1021/bi900101h

Carr SB, Phillips SEV, Thomas CD Structures of replication initiation proteins from staphylococcal antibiotic resistance plasmids reveal protein asymmetry and flexibility are necessary for replication. Nucleic Acids Research 44 2417-2428, 2016
DOI:10.1093/nar/gkv1539
View abstract

Arslan S, Khafizov R, Thomas CD, Chemla YR, Ha T Engineering of a superhelicase through conformational control Science 348 344-347, 2015
DOI:10.1126/science.aaa0445
View abstract

Carr SB, Mecia LB, Phillips SEV, Thomas CD Identification, characterization and preliminary X-ray diffraction analysis of the rolling-circle replication initiator protein from plasmid pSTK1 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 69 1123-1126, 2013
DOI:10.1107/S1744309113023828
View abstract

Zhang WK, Dillingham MS, Thomas CD, Allen S, Roberts CJ, Soultanas P Directional loading and stimulation of PcrA helicase by the replication initiator protein RepD J MOL BIOL 371 336-348, 2007
DOI:10.1016/jmb.2007.05.050

Caryl JA, Thomas CD The pC221 relaxosome: essential interactions for nicking and mobilization. PLASMID 57 195-195, 2007

Espinosa M, Thomas CD Replication - Summary PLASMID 57 183-185, 2007

Zhang W, Dillingham MS, Thomas CD, Allen S, Roberts CJ, Soultanas P Directional loading of the PcrA helicase at the plasmid replication origin oriD by RepD and unwinding of a plasmid. PLASMID 57 195-195, 2007

Zhang W, Dillingham MS, Thomas CD, Allen S, Roberts CJ, Soultanas P Directional loading of the PcrA helicase at the plasmid replication origin oriD by RepD and uniwinding of a plasmid PLASMID 57 188-188, 2007

Thomas CD, Joce CM The inverted complementary repeat is a consequence of, not a requirement for, the action of a replication initiator protein at a replication origin. PLASMID 57 186-186, 2007

Carr SB, Makris G, Phillips SEV, Thomas CD Erratum: Crystallization and preliminary X-ray diffraction analysis of two N-terminal fragments of the DNA-cleavage domain of topoisomerase IV from Staphylococcus aureus (Acta Crystallographica Section F: Structural Biology and Crystallization Communications (2006) F62 (1164-1167)) Acta Crystallographica Section F: Structural Biology and Crystallization Communications 63 59-, 2007
DOI:10.1107/S1744309106054650

Carr SB, Makris G, Phillips SEV, Thomas CD Crystallization and preliminary X-ray diffraction analysis of two N-terminal fragments of the DNA-cleavage domain of topoisomerase IV from Staphylococcus aureus ACTA CRYSTALLOGR F 62 1164-1167, 2006
DOI:10.1107/S1744309106044150

Caryl JA, Thomas CD Investigating the basis of substrate recognition in the pC221 relaxosome MOL MICROBIOL 60 1302-1318, 2006
DOI:10.1111/j.1365-2958.2006.05188.x

Castelletto V, Hamley IW, Kerstens SLH, Deacon S, Thomas CD, Lubbert A, Klok HA Spontaneous condensation in DNA-polystyrene-b-poly(l-lysine) polyelectrolyte block copolymer mixtures EUR PHYS J E 20 1-6, 2006
DOI:10.1140/epje/i2005-10074-x

Caryl JA, Smith M, Thomas CD Reconstitution of a Staphylococcal Plasmid-Protein Relaxation Complex In Vitro Journal of Bacteriology 186 3374-3383, 2004
DOI:10.1128/JB.186.11.3374-3383.2004
View abstract

Smith M, Thomas CD An Accessory Protein Is Required for Relaxosome Formation by Small Staphylococcal Plasmids Journal of Bacteriology 186 3363-3373, 2004
DOI:10.1128/JB.186.11.3363-3373.2004
View abstract

Thomas CD Dimerisation of the RepD initiator protein is a specific and stage-regulated process Plasmid 48 300-301, 2002

Thomas CD, Papadopoulos F, Smith KN The pT181 replicon after 21 years of study: Rolling Circle Plasmid Replication comes of age. Plasmid Biology 2000: International Symposium on Plasmid Molecular Biology. Plasmid 45 pp.140-, 2001

Klimenko D, Convery MA, Rowsell S, Thomas CD, Phillips SEV Crystallization and preliminary X-ray crystallographic studies of RepDC, a hybrid rolling-circle plasmid replication initiator protein Acta Crystallographica. Section D: Biological Crystallography D55 1076-1078, 1999
View abstract

Soultanas P, Dillingham MS, Papadopoulos F, Phillips SEV, Thomas CD, Wigley DB Plasmid replication initiator protein RepD increases the processivity of PcrA DNA helicase Nucleic Acids Research 27 1421-1428, 1999
View abstract

Thomas CD, Nikiforov TT, Connolly BA, Shaw WV Determination of sequence specificity between a plasmid replication initiator protein and the origin of replication Journal of Molecular Biology 254 381-391, 1995
DOI:10.1006/jmbi.1995.0625
View abstract

Thomas CD, Jennings L RepD/D*: A protein-DNA adduct arising during plasmid replication Biochemical Society Transactions 23 pp.442-, 1995

Thomas CD, Modha J, Razzaq TM, Cullis PM, Rivett AJ Controlled high-level expression of the lon gene of Escherichia coli allows overproduction of Lon protease Gene 136 237-242, 1993
DOI:10.1016/0378-1119(93)90471-E
View abstract

Lawson DM, Artymiuk PJ, Yewdall SJ, Smith JMA, Livingstone JC, Treffry A, Luzzago A, Levi S, Arosio P, Cesareni G, Thomas CD, Shaw WV, Harrison PM Solving the structure of human H ferritin by genetically engineering intermolecular crystal contacts Nature 349 541-544, 1991
View abstract

Thomas CD, Balson DF, Shaw WV In vitro studies of the initiation of staphylococcal plasmid replication: Specificity of RepD for its origin (oriD) and characterization of the Rep-ori tyrosyl ester intermediate Journal of Biological Chemistry 265 5519-5530, 1990
View abstract

Catchpole I, Thomas C, Davies A, Dyke KGH The nucleotide sequence of Staphylococcus aureus plasmid pT48 conferring inducible macrolide-lincosamide-streptogramin B resistance and comparison with similar plasmids expressing constitutive resistance Journal of General Microbiology 134 697-709, 1988
View abstract

Thomas CD, Shaw WV, Lawson DM, Treffry A, Artymiuk PJ, Harrison PM Expression and purification of the rat liver ferritin light chain after cloning in Escherichia coli Biochemical Society Transactions 16 838-839, 1988

Thomas CD, Balson DF, Shaw WV Identification of the tyrosine residue involved in bond formation between replication origin and initiator protein of plasmid pC221 Biochemical Society Transactions 16 758-759, 1988