Author/Editor		Plaper, A
Title		Kloniranje in ekspresija cDNA gena, ki se v glivi Rhizopus nigricans inducira v pogojih indukcije hidroksilaznega sistema
Translated title		Cloning and expression of cDNA corresponding to gene induced in Rhizopus nigricans in the conditions of induction of hydroxylase system
Type		monografija
Place		Ljubljana
Publisher		Medicinska fakulteta
Publication year		1995
Volume		str. 104
Language		slo
Abstract		Filamentous fungus Rhizopus nigricans is a suitable model organism for studying of regulation of gene expression by steroids. The fungus has steroid receptors and many proteins are induced by steroids. Among them is a multienzyme system containing cytochrome P-450, which hydroxylates progesterone to 11 alpha-hydroxyprogesterone. With the purpose to study the mechanism of induction of this system on molecular level, we wanted to isolate a gene coding for the cytochrome P-450. We screened a cDNA library prepared in the lambda gt11 expression vector with P-450 antibody probe. A cDNA clone (cDNA of about 500 bp), corresponding to the steroid inducible mRNA was obtained. Positive result in the reaction with affinity purified antibodies from this clone with isolated P-450R.n. was also obtained. In order to determine its function, cDNA of this clone was used for hybridization of a new lamda gt11 cDNA library. The longest cDNA (1,2 kb) obtained by screening was used for subcloning and sequencing. Nucleotide and deduced aminoacid sequence was compared with sequences in the computer database using programs BLAST and FASTA. The comparison revealed no homology with sequences of the cytochrome P-450 gene family but great similarity with aldose-epimerases of numerous organisms. The highest homology was shown with glucose-1-epimerase (mutarotase) from microorganism Acinetobacter calcoaceticus (39.1 pct identity in 361 aminoacid overlap). The deduced protein also contains a sequence typical for the active site of epimerases with histidine and 10 highly Abstract truncated at 3200 characters
Descriptors		RHIZOPUS DNA, COMPLEMENTARY CYTOCHROME P-450 GENE EXPRESSION REGULATION, FUNGAL PROGESTERONE HYDROXYLATION HYDROXYPROGESTERONES GENE LIBRARY ACINETOBACTER CALCOACETICUS ESCHERICHIA COLI RNA RNA, MESSENGER CLONING, MOLECULAR PLASMIDS