By David N. Leff

The sexual revolution really got started back in 1945.

That was the year when molecular biologists Joshua Lederberg and Edward Tatum demonstrated that germs do it.

"They made the mind-blowing discovery," molecular geneticist Frederick Blattner told BioWorld Today, "that bacteria had genes that could be transferred to other microbes; therefore, there was bacterial sex."

For discovering this process of genetic recombination by bacterial conjugation, Lederberg and Tatum shared the 1958 Nobel Prize in physiology or medicine.

The bacterium they stalked, Escherichia coli, has since become, for better or worse, the best-known single cell on earth. The K-12 strain of E. coli in which they studied sexual gene transfer had been found in 1922, infecting a young student at Stanford University who was recovering from diphtheria.

German bacteriologist Theodor Escherich first isolated E. coli in 1885, as an inhabitant of the human colon. There, the germ makes its living as an important member of the intestinal flora, helping to produce the blood-clotting vitamin K and lending a hand in the digestive process.

But as many a tourist, and many a hamburger lover, have learned to their sorrow, E. coli can assault as well as assist that same process. Its pathogenic strains cause traveler's diarrhea and urinary tract infections.

Over 1,600 Genes Hiding Out In Black Boxes

Bullet-shaped E. coli is 10 microns long by 1 wide. The K-12 strain of this subvisible but sophisticated microorganism has a genome that contains 4,639,221 base pairs of DNA. These include 4,288 protein-encoding genes, of which 38 percent have no known function.

Those hefty and long-awaited numbers became known to the world only today, in the current issue of Science, dated Sept. 5, 1997. Blattner, who teaches genetics at the University of Wisconsin, Madison, is the first and principal author of the paper, which announces: "The complete genome sequence of Escherichia coli K-12."

The article's co-authors, from three laboratories in the U.S. and one in Mexico, number 17. It took Blattner and his collaborators 15 years to complete sequencing E. coli's single chromosome. The sequencing proper took off six years ago, with a major grant to Blattner from the National Institutes Of Health's (NIH) National Human Genome Research Institute.

When the project fell short of meeting its 1995 milestone date, the grant was not renewed. The Wisconsin team had only sequenced 1.4 megabases, one-third of the projected total. But out of solidarity with Blattner's dedicated effort, other laboratories declined to bid on a completion grant, so NIH eventually awarded it to him.

On Jan. 16, 1997, having completed 2.6 megabases in 1995, Blattner's group made its final deposit of the E. coli genome to the National Library of Medicine's GenBank. Now the researchers have provided GenBank with a revised, updated definitive version, to coincide with publication of today's report in Science.

Meanwhile in Japan, a rival consortium was completing its own full-length sequencing of the bacterium's genome. But it filed its data with GenBank only on Jan. 23, 1997. "Throughout the project," Blattner said, "there has been friendly, dignified but intense competition with the groups from Japan."

The total cost of his E. coli genome project, Blattner estimated, is $15 million. From this amount he derived a figure of $3 per base, but observed, "In our lab, with a straight face, we can propose to sequence anything for 50 cents a base, even lower for a really big genome with economy of scale."

Now, rather than rest on their genomic laurels, he and his group have taken off in hot pursuit of the E. coli pathogenic mutant that caused the recent recall of 25 million beef patties in the Midwest. "We have under way a project to sequence the E. coli hamburger-disease strain, 015787. As soon as we finished the K-12 genome," he said, "we started on the hamburger. We've been trying to get grant money for this project for the past four years. Our work on it is far along, and going very nicely."

This deadly strain, he pointed out, is "not the same one that causes traveler's diarrhea, but similar. Other E. coli do that. 0151787 has a much more virulent phenotype. If you get the worst form of it, it basically decimates the kidneys and kills you." There were 8,000 cases two years ago, he noted, with a 5 percent mortality rate.

Another front-burner project is to assign functions to the nearly 40 percent of E. coli's genes that remain total mysteries. One approach, Blattner described, "is a program to measure the expression of every gene in defined conditions. Another is to produce conditional mutations in knockout genes.

In the late 1970s, Eli Lilly and Co., of Indianapolis, cloned the human gene for insulin in a weakened strain of K-12, then filed a new drug application with the FDA on May 14, 1982. Yet other host cells have since largely edged out E. coli as a recombinant host organism.

Blattner commented: "Now that we've got the complete genome sequence, it would be very interesting to try to use this information to improve it, for production purposes. One obvious thing is getting rid of the genes that are not necessary. But there's also ideas for enhancement, to re-engineer this, based on the knowledge we have, so that it becomes a better tool of production." *

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