TY - JOUR
T1 - Identification of proteins and protein domains that contact DNA within adenovirus nucleoprotein cores by ultraviolet light crosslinking of oligonucleotides 32P-labelled in vivo
AU - Chatterjee, Pradeep K.
AU - Vayda, Michael E.
AU - Flint, S. J.
N1 - Funding Information:
LVe thank Rhonda Lunt and Margifl Young tbl excellent technical assistance. and Ueng-Cheng Yang for many helpful discussions. This work was supported by a, grant (NP 464-F) from the American Cancer Society.
PY - 1986/3/5
Y1 - 1986/3/5
N2 - A new approach to the identification of DNA binding proteins has been developed and used to study the DNA-protein interactions within the nucleoprotein core of subgroup C adenoviruses. Virions labelled in vivo with [32P]orthophosphate were exposed to ultraviolet light and the DNA digested by chemical or enzymatic methods. Labelled phosphoamino acids of the virion phosphoproteins were selectively hydrolysed by alkali, permitting proteins crosslinked to DNA to be identified by virtue of their covalently attached, 32P-labelled nucleotides. In parallel experiments, [3H]arginine-labelled virions were crosslinked by exposure to ultraviolet light and analysed by more conventional methods. The results indicate that proteins VII and V lie in close contact with viral DNA within the core. The compact arrangement of the nucleoprotein core appears to be capable of trapping protein VII molecules that are not covalently attached to DNA after exposure to ultraviolet light, suggesting that viral DNA might be wrapped around clusters of protein VII molecules. The domains of protein VII that lie in contact with DNA were identified by partial proteolytic mapping of the sites of covalent-attachment of the 32P-labelled oligonucleotides. The implications of these data for the nature of the interactions that mediate the packaging of viral DNA within the nucleoprotein core of adenovirions are discussed.
AB - A new approach to the identification of DNA binding proteins has been developed and used to study the DNA-protein interactions within the nucleoprotein core of subgroup C adenoviruses. Virions labelled in vivo with [32P]orthophosphate were exposed to ultraviolet light and the DNA digested by chemical or enzymatic methods. Labelled phosphoamino acids of the virion phosphoproteins were selectively hydrolysed by alkali, permitting proteins crosslinked to DNA to be identified by virtue of their covalently attached, 32P-labelled nucleotides. In parallel experiments, [3H]arginine-labelled virions were crosslinked by exposure to ultraviolet light and analysed by more conventional methods. The results indicate that proteins VII and V lie in close contact with viral DNA within the core. The compact arrangement of the nucleoprotein core appears to be capable of trapping protein VII molecules that are not covalently attached to DNA after exposure to ultraviolet light, suggesting that viral DNA might be wrapped around clusters of protein VII molecules. The domains of protein VII that lie in contact with DNA were identified by partial proteolytic mapping of the sites of covalent-attachment of the 32P-labelled oligonucleotides. The implications of these data for the nature of the interactions that mediate the packaging of viral DNA within the nucleoprotein core of adenovirions are discussed.
UR - http://www.scopus.com/inward/record.url?scp=0023041995&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0023041995&partnerID=8YFLogxK
U2 - 10.1016/0022-2836(86)90477-8
DO - 10.1016/0022-2836(86)90477-8
M3 - Article
C2 - 3712442
AN - SCOPUS:0023041995
SN - 0022-2836
VL - 188
SP - 23
EP - 37
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 1
ER -