The human intestinal system is a battlefield between the “good” bacteria that, for example, help with digestion and the “bad” bacteria that can make us sick. One weapon the body uses to kill the bad bacteria is antimicrobial peptides, which are small protein fragments. However, the body produces enzymes that split the antimicrobial peptides into hundreds of even smaller fragments. Researchers had thought that this process destroyed the weapon, but new research shows that each small fragment specifically attacks and kills especially bad bacteria. The researchers will now develop this naturally occurring fragmentation bomb to provide an alternative to antibiotics and to improve general intestinal health.
The task sounds simple enough: fight the enemy and support the allies. But understanding these processes can be difficult when this struggle takes place in the innermost part of our intestinal system. Nevertheless, understanding the defence mechanism in people’s gut is extremely important because it is key to avoiding disease and strengthening health. Now researchers have made a major breakthrough in understanding how the intestinal system keeps external enemies from invading the body.
“The Paneth cells that line the small intestine secrete defensins: small protein fragments that can kill pathogenic bacteria. We did not understand why the intestinal fluid itself seems to split some of these defensins into hundreds of smaller fragments. Now we can see that this process does actually does not destroy the defensins; on the contrary, it creates hundreds of new and more specific weapons. Now we hope to copy this fragmentation bomb to treat bacterial infections – as an alternative to the antibiotics we have today that are increasingly ineffective,” explains a researcher behind the study, Benjamin Anderschou Holbech Jensen, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen.
Skewed bacterial balance
To understand why the defensive weapons secreted by the Paneth cells of the small intestine were apparently neutralized as soon as they entered the intestine, the researchers decided to examine how the enzymes –proteases – in the duodenal fluid of the small intestine would react in the laboratory with two of the most important small intestinal defensins: human α-defensin 5 (HD-5) and 6 (HD-6).
“We used liquid chromatography and mass spectrometry to monitor how the enzymes fragmented the defensins. This enabled us to both identify each defensin fragment and test how they affected the growth of different types of bacteria. The proteases apparently did not affect HD-6 at all due to the formation of a protective peptide nanonet shielding this particular defensin from the enzymatic degradation, whereas HD-5 was split into hundreds of small fragment combinations. The scientific community at large, expected that this would inactivate its antimicrobial properties. So we were surprised when we tested how the fragments affected different bacteria.”
The researchers had expected that the HD-5 fragments would be wholly or partly ineffective in retarding the growth of bacteria. Instead, the hundreds of fragments produced new and even more active combinations, and the researchers found that HD-5 positively affected the bacterial balance.
“The pathogens died off, and even more favourable conditions were created for the bacteria that are desirable in the gut. So a single simple peptide, HD-5, functions as a kind of fragmentation bomb that seems to be key in fine-tuning our intestinal system and providing a healthy balance.”
Promising pathways for developing drugs
The new finding is surprising because researchers had thought that biological degradation inactivates one of the intestine’s most important weapons against external enemies. Instead, the degradation process leads to hundreds of fragments, each of which has unique specificity and a strategy for eliminating bacteria.
“We sensed that HD-5 in particular and defensins in general must be important, since evolution has preserved them for millennia and across different species, including plants, fish, birds and mammals – with very little variation between species. They are thus one of the most frequently conserved defence mechanisms in biology. We are extremely fascinated that evolution has actually developed clusters of fragments that are further activated when they reach the intestine and whose cocktail is so targeted and effective against bad bacteria – without damaging the good bacteria.”
This discovery may prove very valuable in fighting the multidrug-resistant bacteria that existing antibiotics cannot eliminate. The really good news is that, although HD-5 is a relatively long protein fragment of 34 amino acids that strongly depends on an extremely specific structure that is difficult to produce synthetically, producing the typical nine-peptide-long linear microfragments that HD-5 becomes when split would be relatively simple.
“These results significantly expand our understanding of how the body maintains balance in our microbiome and how complex the intestinal defence mechanisms are. They also show extremely promising avenues for developing drugs to combat not only bacterial infections but also diabetes, cardiovascular diseases and other complications related to the microbiome since our other experiments show that the defensins significantly lower cholesterol levels and, at least in mice, can reduce the risk of developing nonalcoholic fatty liver disease and also treat it. We are currently expanding these studies to specific human models of diabetes-accelerated arteriosclerosis and general atherosclerosis, which cause nearly one third of all deaths in high-income countries.”
“Paneth cell α-defensins HD-5 and HD-6 display differential degradation into active antimicrobial fragments” has been published in Proceedings of the National Academy of Sciences of the United States of America. “Human Paneth cell α-defensin 5 treatment reverses dyslipidemia and improves glucoregulatory capacity in diet-induced obese mice” has been published in the American Journal of Physiology, Endocrinology and Metabolism. In 2017, the Novo Nordisk Foundation awarded a grant to Benjamin Anderschou Holbech Jensen for the project A Novel Oral Combination Therapy Targeting the Gut Microbiome to Alleviate Insulin Resistance and Type 2 Diabetes–linked Aortic Valve Stenosis.