A.J.J. van Ooyen

Bio: A.J.J. van Ooyen is an academic researcher from University of Zurich. The author has contributed to research in topics: Gene & Transcription (biology). The author has an hindex of 10, co-authored 12 publications receiving 1073 citations. Previous affiliations of A.J.J. van Ooyen include Wageningen University and Research Centre & DSM.

Kluyveromyces as a host for heterologous gene expression: expression and secretion of prochymosin.

Abstract: We have developed the yeast Kluyveromyces lactis as a host organism for the production of the milk-clotting enzyme chymosin. In contrast to Saccharomyces cerevisiae, we found that this yeast is capable of the synthesis and secretion of fully active prochymosin. Various signal sequences could be used to efficiently direct the secretion of prochymosin in Kluyveromyces, but not in S. cerevisiae. We conclude that the efficient synthetic and secretory capacity of this heterologous protein is a property of the yeast Kluyveromyces. These results have led to the development of a large scale production process for chymosin.

Comparison of total sequence of a cloned rabbit beta-globin gene and its flanking regions with a homologous mouse sequence.

Abstract: The nucleotide sequence of a cloned rabbit chromosomal DNA segment of 1620 nucleotides length which contains a beta-globin gene is presented. The coding regions are separated into three blocks by two intervening sequences of 126 and 573 base pairs, respectively. The rabbit sequence was compared with a homologous mouse sequence. The segments flanking the rabbit gene, as well as the coding regions, the 5' noncoding and part of the 3' noncoding messenger RNA sequences are similar to those of the mouse gene; the homologous introns, despite identical location, are distinctly dissimilar except for the junction regions. Homologous introns may be derived from common ancestral introns by large insertions and deletions rather than be multiple point mutations.

Subfragments of the large terminal repeat cause glucocorticoid-responsive expression of mouse mammary tumor virus and of an adjacent gene

Abstract: After transfection of mouse mammary tumor virus (MMTV) proviral DNA into cultured cells, the DNA is transcribed in a glucocorticoid-sensitive fashion. The large terminal repeat (LTR) region of MMTV is 1,328 nucleotides long and contains the regulatory information necessary for the hormonal response. We have constructed a MMTV LTR-thymidine kinase (tk) chimeric gene and have tested the biological activity of molecules containing various deletions in the LTR after transformation of LTK- APRT- mouse cells. In the TK+ transformants, both a LTR- tk chimeric RNA and an authentic tk RNA are correctly initiated and transcribed. The synthesis of the chimeric RNA as well as that of the tk RNA is hormonally regulated. A plasmid containing 202 nucleotides of LTR DNA 5' to the RNA initiation site is fully sensitive to glucocorticoids; 50 nucleotides still cause a residual inducibility.

DNA sequences preceding the rabbit beta-globin gene are required for formation in mouse L cells of beta-globin RNA with the correct 5' terminus

Abstract: Mouse thymidine kinase-negative (TK-) L cells were transformed with concatenates of cloned herpes simplex virus 1 TK DNA and different rabbit beta-globin DNAs in which the globin genes were preceded by native flanking sequences of 14, 66, 76, 425, and 1500 nucleotides.l In all cases, selection for TK+ cell lines led to a high yield of lines producing 5-1500 mature rabbit beta-globin-specific RNA strands per cell. The 5' termini of the transcripts mapped to (i) the "cap" nucleotide, (ii) positions 42 to 48 nucleotides downstream from the cap site, or (iii) positions in the vector DNA preceding the gene. In the case of the gene with only 14 base pairs of 5' flanking sequence, a high level of rabbit beta-globin RNA was produced, but none of the transcripts had the correct 5' end; most of them originated in the vector moiety. With 66 base pairs of 5' flanking sequence, 5% of the 5' termini were correct, and with 76 or more base pairs, 30-85% were correct. The region between 14 and 66 base pairs preceding the cap site contains the Hogness box and appears to be essential for correct initiation of transcription. The region between 66 and 76 base pairs before the cap site contains a variant of the canonical sequence G-G-C-T-C-A-A-T-C-T found preceding many other genes at a similar location, and this region may modulate the efficiency of transcription. The sequence of 425 nucleotides preceding the rabbit beta-globin gene is reported.

A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences.

Abstract: Some simple formulae were obtained which enable us to estimate evolutionary distances in terms of the number of nucleotide substitutions (and, also, the evolutionary rates when the divergence times are known). In comparing a pair of nucleotide sequences, we distinguish two types of differences; if homologous sites are occupied by different nucleotide bases but both are purines or both pyrimidines, the difference is called type I (or “transition” type), while, if one of the two is a purine and the other is a pyrimidine, the difference is called type II (or “transversion” type). Letting P and Q be respectively the fractions of nucleotide sites showing type I and type II differences between two sequences compared, then the evolutionary distance per site is K = — (1/2) ln {(1 — 2P — Q) }. The evolutionary rate per year is then given by k = K/(2T), where T is the time since the divergence of the two sequences. If only the third codon positions are compared, the synonymous component of the evolutionary base substitutions per site is estimated by K'S = — (1/2) ln (1 — 2P — Q). Also, formulae for standard errors were obtained. Some examples were worked out using reported globin sequences to show that synonymous substitutions occur at much higher rates than amino acid-altering substitutions in evolution.