The point mutation M26 in the ade6 gene of Schizosaccharomyces pombe increases recombination frequency by an order of magnitude in comparison with other mutations in the same gene. The hypothesis is tested that this hot spot of recombination requires a specific nucleotide sequence at the M26 site. The DNA sequence is altered systematically by in vitro mutagenesis, and the resulting sequences are introduced into the ade6 gene in vivo by gene replacement. It results that any change of the heptanucleotide ATGACGT leads to loss of high frequency of recombination. Thus this oligonucleotide sequence is necessary for high frequency of recombination, but it seems not to be sufficient.

Glycosylatedcellulases from Celhdomonas jimi were compared with their non-glycosylated counterparts synthesized in Escherichia coli from recombinant DNA. Glycosylation of the enzymes does not significantly affect their kinetic properties, or their stabilities towards heat and pH. However, the glycosylated enzymes are protected from attack by a C.fimi protease when bound to cellulose, while the non-glycosylated enzymes yield active, truncated products with greatly reduced affinity for cellulose.

A sensitive immunoassay developed previously was used to isolate a series of Escherichia coli clones expressing cellulase genes from Cellulomonas $mi. The clones fell into three groups. Clones in the first group contained plasmids with a 6.6 kb insert of C.$mi DNA, were strongly antigenic, and contained low levels of CM-cellulase activity. Those in the second group contained plasmids with a 5-0 kb insert, were weakly antigenic, and contained high levels of CM-cellulase activity. Those in the third group contained plasmids with a 5.6 kb insert, were weakly antigenic, and contained low levels of CM-cellulase activity. Restriction analysis showed that the three groups carried different cellulase genes. All of these CM-cellulase activities were exported to the periplasm in E. coli, but with different efficiencies. These activities represented important components of the C. $mi cellulase complex. Their properties indicated that they were different from each other and that they probably had complementary actions.

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Cellulomonas fimi produces an endoglucanase and an exoglucanase which bind strongly to cellulose. Each enzyme contains three distinct regions: a short sequence of about 20 amino acids containing only proline and threonine (the Pro-Thr box); an irregular region, rich in hydroxyamino acids, of low charge density, and which is predicted to have little secondary structure; and an ordered region of higher charge density which contains a potential active site, and which is predicted to have secondary structure. The Pro-Thr box is conserved almost perfectly in the two enzymes. The irregular regions are 50% conserved, and the conserved sequences include four Asn-Xaa-Ser/Thr sites. The ordered regions appear not to be conserved, but the potential active sites both have the sequence Glu-Xaa7-Asn-Xaa6-Thr; they occur at widely separated sites in the two regions. The order of the regions is reversed in the two enzymes: irregular-Pro-Thr box-ordered in the endoglucanase; ordered-Pro-Thr box-irregular in the exoglucanase. The genes for the two enzymes appear to have arisen by shuffling of two conserved sequences and either one or two other sequences.