Cloning and characterization of the three enzyme structural genes QUTB, QUTC and QUTE from the quinic acid utilization gene cluster in Aspergillus nidulans

1985 ◽  
Vol 9 (4) ◽  
pp. 305-311 ◽  
Author(s):  
Alastair R. Hawkins ◽  
Antonio J. Da Silva Francisco ◽  
Clive F. Roberts
1990 ◽  
Vol 265 (2) ◽  
pp. 337-342 ◽  
Author(s):  
R K Beri ◽  
S Grant ◽  
C F Roberts ◽  
M Smith ◽  
A R Hawkins

The three enzymes necessary to catabolize quinate to protocatechuate are inducible by quinic acid, and transcription of their corresponding genes is controlled by the action of a positively acting activator gene and a negatively acting repressor gene. Transformed strains of Aspergillus nidulans containing multiple copies of the activator gene (QUTA) but single copies of the other QUT genes retain normal regulation of the gene cluster and do not show any overexpression of the three quinic acid catabolic enzymes. Transformed strains containing equal multiple copies of the activator gene (QUTA) and QUTE (encoding catabolic 3-dehydroquinase), but single copies of the other QUT genes, retain normal regulation of the QUT gene cluster, but selectively overexpress the QUTE gene upon quinic acid induction. Data are presented that strongly suggested that the gene QUTG, which is physically located within the QUT gene cluster and for which no function has been identified, is not required for expression of the gene cluster and does not encode a chlorogenic acid esterase.


1993 ◽  
Vol 296 (2) ◽  
pp. 451-457 ◽  
Author(s):  
A R Hawkins ◽  
J D Moore ◽  
A M Adeokun

The AROM protein of Aspergillus nidulans is a multidomain pentafunctional polypeptide that is active as a dimer and catalyses steps 2-6 in the prechorismate section of the shikimate pathway. The three C-terminal domains (including the type I 3-dehydroquinase) of the AROM protein are homologous with the qutR-encoded QUTR protein that represses transcription of the eight genes comprising the quinic acid utilization (qut) gene cluster, and the two N-terminal domains are homologous with the qutA-encoded QUTA protein that transcribes the qut genes. As part of a larger research programme designed to compare the structures of the three proteins and to probe the domain structure and interaction within each protein, we have overproduced and purified the 3-dehydroquinase domain of the AROM protein. Additionally we have overproduced and purified the qutB-encoded quinate dehydrogenase and overproduced the qa-2 encoded type II 3-dehydroquinase of Neurospora crassa. We report that the AROM 3-dehydroquinase domain has a monomeric native state, with an apparent kcat./Km ratio that is approx. 160-fold lower than the value for the native N. crassa AROM protein. The AROM protein 3-dehydroquinase domain is sensitive to inactivation by borohydride in the presence of the substrate 3-dehydroquinate, confirming that it is a typical type I 3-dehydroquinase. The purified quinate dehydrogenase is bifunctional, being able to metabolize shikimate as a substrate. The apparent Km values for quinate (450 microM), shikimate (1.7 mM) and NAD+ (150 microM) are all similar to values reported for the qa-3-encoded enzyme from N. crassa.


1988 ◽  
Vol 214 (2) ◽  
pp. 224-231 ◽  
Author(s):  
Alastair R. Hawkins ◽  
Heather K. Lamb ◽  
Melanie Smith ◽  
John W. Keyte ◽  
Clive F. Roberts

Microbiology ◽  
1988 ◽  
Vol 134 (2) ◽  
pp. 347-358 ◽  
Author(s):  
S. GRANT ◽  
C. F. ROBERTS ◽  
H. LAMB ◽  
M. STOUT ◽  
A. R. HAWKINS

PLoS ONE ◽  
2012 ◽  
Vol 7 (4) ◽  
pp. e35450 ◽  
Author(s):  
Kirsi Bromann ◽  
Mervi Toivari ◽  
Kaarina Viljanen ◽  
Anu Vuoristo ◽  
Laura Ruohonen ◽  
...  

1990 ◽  
Vol 223 (1) ◽  
pp. 17-23 ◽  
Author(s):  
Heather K. Lamb ◽  
Alastair R. Hawkins ◽  
Melanie Smith ◽  
Ian J. Harvey ◽  
John Brown ◽  
...  

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