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Stephen ArnoffJon K. de Riel, PhD

 

Assistant Professor, Fels Institute for Cancer Research

and Molecular Biology

Telephone:  215-707-4347

Fax:  215-707-5963

Email: jon.deriel@temple.edu

 

Fels Institute for Cancer Research and Molecular Biology

 

Educational Background:

 

Harvard Medical School, Boston, MA

 

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Research Interests:

 

Our longstanding collaboration with Dr. Earl Henderson (Fels Institute) is focused on the molecular biology of the denV gene from bacteriophage T4, which encodes endonuclease V, a 16 kDa DNA repair protein that catalyzes two distinct steps in the base excision repair of pyrimidine dimers. Since 1984, we have extensively characterized the biology of den V-mediated UV repair in both prokaryotic and eukaryotic cells. Because endonuclease V possesses two distinct catalytic activities (apyrimidine dimer-DNA glycosylase activity and an apyrimidinic-site endonuclease) in a single small structural domain, its mechanism is of considerable interest. For structure-function analysis of the protein, we have developed a method of random targeted mutagenesis, based on a previously published procedure for polymerase chain reaction mutagenesis that can be used to blanket a selected small region of the gene with random mutation. The resulting mutations are then identified by sequencing and correlated with the catalytic properties of the corresponding altered proteins. The method is useful because it allows random targeted mutagenesis to be performed directly in a plasmid expression vector, permitting immediate functional screening of the resulting mutants. This procedure has been used to analyze the functional importance of the Ala(116)-Lys(121) region of endonuclease V. Screening of the resulting clones detected a high percentage of mutants (19/22) which included 15 unique mutants. Analysis of the UV-repair properties of these mutants, and studies of purified endonuclease V from them, show an important role for this region in substrate binding, and also suggest a direct correlation between UV survival and both DNA binding and pyrimidine dimer-DNA glycosylase activity, but not apyrimidinic-site endonuclease activity. Current efforts are concerned with extending random targeted mutagenesis studies to other regions of the molecule, with the intent of separating the two catalytic activities into two different modified proteins; and also examining the effects of the den V gene on mutagenesis in human cell lines.

 

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PUBMED PUBLICATIONS :


Recent Medically Related Publications, Obtained from PubMed (Click on PubMed ID to view abstract)

15558027. Kaur GP, Reddy DE, Zimonjic DB, de Riel JK, Athwal RS, Functional identification of a BAC clone from 16q24 carrying a senescence gene SEN16 for breast cancer cells. Oncogene 24:1(47-54)2005 Jan 6

11580279. Li J, Huang P, Chen C, de Riel JK, Weinstein H, Liu-Chen LY, Constitutive activation of the mu opioid receptor by mutation of D3.49(164), but not D3.32(147): D3.49(164) is critical for stabilization of the inactive form of the receptor and for its expression. Biochemistry 40:40(12039-50)2001 Oct 9

11434771. Xu W, Li J, Chen C, Huang P, Weinstein H, Javitch JA, Shi L, de Riel JK, Liu-Chen LY, Comparison of the amino acid residues in the sixth transmembrane domains accessible in the binding-site crevices of mu, delta, and kappa opioid receptors. Biochemistry 40:27(8018-29)2001 Jul 10

11076532. Xu W, Chen C, Huang P, Li J, de Riel JK, Javitch JA, Liu-Chen LY, The conserved cysteine 7.38 residue is differentially accessible in the binding-site crevices of the mu, delta, and kappa opioid receptors. Biochemistry 39:45(13904-15)2000 Nov 14

10416823. Li JG, Chen C, Yin J, Rice K, Zhang Y, Matecka D, de Riel JK, DesJarlais RL, Liu-Chen LY, ASP147 in the third transmembrane helix of the rat mu opioid receptor forms ion-pairing with morphine and naltrexone. Life Sci 65:2(175-85)1999

10214970. Xu W, Ozdener F, Li JG, Chen C, de Riel JK, Weinstein H, Liu-Chen LY, Functional role of the spatial proximity of Asp114(2.50) in TMH 2 and Asn332(7.49) in TMH 7 of the mu opioid receptor. FEBS Lett 447:2-3(318-24)1999 Mar 26

8947926. Chen XH, Liu-Chen LY, Tallarida RJ, Geller EB, de Riel JK, Adler MW, Use of a mu-antisense oligodeoxynucleotide as a mu opioid receptor noncompetitive antagonist in vivo. Neurochem Res 21:11(1363-8)1996 Nov

8542317. Chen XH, Geller EB, de Riel JK, Liu-Chen LY, Adler MW, Antisense oligodeoxynucleotides against mu- or kappa-opioid receptors block agonist-induced body temperature changes in rats. Brain Res 688:1-2(237-41)1995 Aug 7

7768887. Xue JC, Chen C, Zhu J, Kunapuli SP, de Riel JK, Yu L, Liu-Chen LY, The third extracellular loop of the mu opioid receptor is important for agonist selectivity. J Biol Chem 270:22(12977-9)1995 Jun 2

8316218. Mauro DJ, De Riel JK, Tallarida RJ, Sirover MA, Mechanisms of excision of 5-fluorouracil by uracil DNA glycosylase in normal human cells. Mol Pharmacol 43:6(854-7)1993 Jun

2785906. Ghezzo F, Valpreda S, De Riel JK, Baserga R, Identification of serum-responsive elements in the promoter of human calcyclin, a growth-regulated gene. DNA 8:3(171-7)1989 Apr

2927423. Henderson EE, Valerie K, Green AP, de Riel JK, Host cell reactivation of CAT-expression vectors as a method to assay for cloned DNA-repair genes. Mutat Res 220:2-3(151-60)1989 Mar-May

3308908. Kaczmarek L, Calabretta B, Ferrari S, de Riel JK, Cell-cycle-dependent expression of human ornithine decarboxylase. J Cell Physiol 132:3(545-51)1987 Sep

2446955. Valerie K, Fronko G, Long W, Henderson EE, Nilsson B, UhlÚn M, de Riel JK, Production and detection of coliphage T4 endonuclease V polyclonal and monoclonal antibodies using staphylococcal protein-A hybrid proteins. Gene 58:1(99-107)1987

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