Biomedical subjects

T G Guilliams

Publications and source records attributed to T G Guilliams.

3 recordsLinked to original sources

PubMed1997

Site directed DNA joining.

We have previously identified a cDNA encoding part of the amino terminal portion of the large subunit of replication factor c (RF-C, AP-1), which we have named VDJP. Analysis of VDJP demonstrated that it has amino acid homology to bacterial DNA ligases and specific binding to the nonamer portion of the V(D)J recombination signal sequence motif. In this report, we demonstrate that VDJP is capable of forming a covalent bond between DNA fragments in a sequence dependent fashion. The VDJP mediated DNA ligation reaction is neither dependent on the presence of compatible DNA ends nor on sequence homology between the DNA fragments that are joined. Furthermore, we show that the covalent junction between the DNA fragments is resistant to proteases and phenol, and therefore not protein linked.

DNA Ligases↗

PubMed1995

Cloning of the murine cDNA encoding VDJP, a protein homologous to the large subunit of replication factor C and bacterial DNA ligases.

A putative full-length 1.7-kb cDNA, encoding a murine protein that specifically binds to the nonamer portion of the V(D)J recombinational signal sequence (RSS) element, has been cloned. By its sequence analysis, this cDNA is identical to a portion of the 4.5-kb murine replication factor C large-subunit-encoding cDNA. By Northern blot analysis, the 1.7-kb mRNA species is observed in murine immature B cells but not in non-lymphoid cells and tissues, while the 4.5-kb replication factor C-encoding cDNA is expressed in all cell types. The deduced VDJP amino-acid sequence includes a region of homology with bacterial DNA ligases at the C terminus of each of the proteins. VDJP has been synthesized as a fusion protein in bacteria, and the purified protein has been previously shown to mediate the joining of DNA fragments in a V(D)J RSS-dependent fashion (Guilliams et al., Biochem. Biophys. Res. Commun. 202 (1994) 1134-1141).

Amino Acid Sequence↗

PubMed1994

Distance and end configuration effects on VDJP-mediated DNA joining.

We have previously reported the cloning of a protein, VDJP, that is capable of binding the nonamer element of the V(D)J Recombinational Signal Sequence (RSS) as well as joining linear DNA fragments containing RSS elements in vitro. We show here that the linearized DNA molecules must contain a 5' extension or blunt end in order to be joined by VDJP. DNAs with 3' extensions are not efficiently joined by VDJP. Furthermore, the joining activity of DNAs with 5' extensions is significantly increased as the distance between the end and the RSS decreases. It is not yet clear what role VDJP plays in vivo, because our assay may not mimic exactly the in vivo DNA intermediates.

Animals↗