Identification of an attenuating region in the bovine follicle-stimulatinghormone beta subunit mRNA that decreases its expression in E-coli

Follicle-stimulating hormone (FSH) is one of the key regulators of gonadal function in mammals. Recombinant DNA expression of this hormone has proved to be a difficult task as expression levels are invariably low, irrespectiv e of the expression system employed. In the present study, we have attempte d to identify reasons for this low expression using bacterial expression ve ctors, and we report here the identification of a theoretically predicted h airpin structure in the mRNA corresponding to the N-terminal portion of the mature coding portion of bFSH beta cDNA that is responsible for attenuatin g its expression in E. coli. When full-length FSH beta was expressed using the bacterial expression vector, a very low expression was obtained. Howeve r, when fragments of FSH beta with N-terminal deletions (amino acids 24-110 and 13-110) were expressed using the same expression strategy, a 30- to 40 -fold higher expression was observed. This low expression of FSH beta could be attributed to a hairpin structure present in the first 12 codons of mat ure FSH beta mRNA. Disruption of this structure without changing the amino acid sequence resulted in a higher level of expression of FSH beta. The pre dicted hairpin structure, though away from the transcriptional and translat ional start site, was able to downregulate the expression of FSH beta proba bly by impeding the movement of ribosomes. (C) 1999 Published by Elsevier S cience B.V. All rights reserved.