St. Louis, (Date) -- The bacterium that causes most peptic ulcers clings to the stomach wall by locking onto specific receptors on cells in the gastric lining. Collaborators in Sweden and the United States now have identified and isolated a bacterial protein that allows the microbe to cling.
"With this attachment protein in hand, it eventually should be possible to develop a vaccine against peptic ulcers and gastric cancer," says Thomas Borén, D.D.S., Ph.D., an assistant professor of oral biology at Umeå University in Sweden. Borén headed the research team, which published its findings in the Jan. 16 issue of Science.
When Borén was at Washington University School of Medicine in St. Louis in 1993, he discovered that Helicobacter pylori, the bacterium that infects the gastric lining, attaches to a carbohydrate called the Lewis b (Leb) blood group antigen. This antigen is found on the red blood cells of people with type O blood, but it also occurs on the gastric lining. "So the bacterium prefers to bind to receptors present in blood group O individuals," Borén says. "And we have found that this binding property is highly associated with H. pylori isolated from patients with ulcer disease."
Most of the world's population is infected with H. pylori, though the prevalence is higher in developing countries and in regions of poverty in the Western world. The bacterium probably is spread by unclean fingers and hands and contaminated food and water. More than one-third of Americans have this infection, and about 20 percent of them eventually develop peptic ulcer disease. Most gastric cancers also occur in people with current or past H. pylori infections. Although stomach cancer is rare in the United States, it is the most common male cancer in some parts of the world.
"So there's a real need for new ways to prevent or cure H. pylori infection," says Douglas E. Berg, Ph.D., the American member of the research team. "In places where people are at high risk for infection, such as shanty towns in developing countries, reinfection after curing with antibiotics is commonplace. So an effective, cheap vaccine that boosted immunity after every exposure would be incredibly important." Berg is the Alumni Professor in Molecular Microbiology and a professor of genetics at Washington University School of Medicine in St. Louis.
In the study, the collaborators focused on the microbial part of the interaction between H. pylori and Leb -- the second half of the Velcro sandwich. The Swedish group showed that a protein they located on the surface of H. pylori -- BabA (blood group antigen binding adhesin) -- attaches to the Leb antigen. They also found that H. pylori strains from different parts of the world had adhesins that were the same size and had the same initial 20 amino acids. "Biologically conserved bacterial surface proteins have great potential as vaccine candidates," Borén says.
If part of the BabA molecule could be used as a vaccine, it would prime the immune system to attack similar proteins, preventing H. pylori from clinging to the gastric lining. Dislodged as the stomach churns or empties, the bacterium would be less able to deliver toxic products to human tissue and therefore less likely to cause disease.
To find the BabA protein, Borén's group invented a clever technique called ReTagging (Receptor Activity Directed Affinity Tagging). First, the researchers attached several Leb molecules to a large protein called albumin, creating a structure that looked like a potato stuck with toothpicks. Then they added a Y-shaped "toothpick" with biotin (actually a vitamin) on one tip. By mixing these molecular potatoes with an H. pylori suspension and exposing the mixture to ultraviolet light, they chemically linked the Y toothpick to whatever had bound to their synthetic Leb receptor. Using special chemical reagents, they then snapped the Y toothpick off the potato, leaving part of it stuck to the bacterium. In this way, they added a biotin label to the protein that had bound to Leb.
To isolate this protein, the researchers ground up the tagged H. pylori and added magnetic beads that had been coated with avidin, a protein that binds tenaciously to biotin. The avidin attracted the biotin out of the mixture along with the sought-after protein. After pulling out the beads with a magnet, the researchers removed the bacterial protein.
After determining part of the amino acid sequence of this protein, Borén's group deduced the type of DNA sequence that could encode it. The researchers then chemically synthesized pieces of DNA with that sequence to serve as a primer in a powerful technique called PCR (the polymerase chain reaction). This allowed them to recover the entire gene from the bacterium. But instead of finding one such gene, they found a complex family, which included two closely related members.
To determine which gene was functional, one of Borén's students, Johan Ögren, spent the spring of 1997 in Berg's lab in St. Louis. He and Dangeruta Kersulyte, a research associate in molecular microbiology at Washington University School of Medicine, inactivated each of the two related genes to see which one was critical for adhesion. Only the loss of the gene called babA2 prevented the bacterium from binding to Leb.
"We propose that the protein product of this gene, BabA, allows H. pylori to stick to the gastric epithelium and play a critical role in the efficient delivery of factors that directly damage gastric tissues," Borén says.
Thomas Borén can be reached at: Thomas.Boren@oralbio.umu.se
Douglas Berg can be reached at: 314-286-0119
The work was supported by grants from the Swedish Society of Medicine, Lion's Cancer Research Foundation, Umeå University, the Magnus Bergvall Foundation, the Swedish Medical Research Council, the Swedish Society for Medical Research, the Royal Swedish Academy of Sciences, the Umeå University-Washington University Scientific Exchange Program, the National Institutes of Health and the American Cancer Society.
Ilver D, Arnqvist A, Ögren J, Frick I-M, Kersulyte D, Incecik ET, Berg DE,
Covacci A, Engstrand L, Borén T.
Helicobacter pylori Adhesin Binding Fucosylated Histo-Blood Group Antigens
Revealed by ReTagging.
Science, vol. 279, Jan. 16, 1998.
Journal
Science