Microbial metalloproteomes—the set of proteins that have metal-binding capacity by either being metalloproteins or having metal-binding sites—are much more extensive and diverse than previously recognized, and promise to provide key insights for cell biology, microbial growth and toxicity mechanisms, according to a new study published online 18 July in the journal Nature.

Researchers from the US Department of Energy’s Lawrence Berkeley National Laboratory in collaboration with scientists at the University of Georgia developed a robust, metal-based approach to determine all metals an organism assimilates and identify its metalloproteins on a genome-wide scale. Their findings suggest that the microbial world features a broader and more diverse array of metal-driven chemical processes than previously imagined. This could lead to new ways to harness metal-driven chemical processes to create next-generation biofuels or to clean up environmental contaminants.

Metal ion cofactors afford proteins virtually unlimited catalytic potential, enable electron transfer reactions and have a great impact on protein stability. Consequently, metalloproteins have key roles in most biological processes, including respiration (iron and copper), photosynthesis (manganese) and drug metabolism (iron). Yet, predicting from genome sequence the numbers and types of metal an organism assimilates from its environment or uses in its metalloproteome is currently impossible because metal coordination sites are diverse and poorly recognized.

—Cvetkovic et al.

The new metal-based approach, the team says, “shifts the focus from classical protein-based purification to metal-based identification and purification by liquid chromatography, high-throughput tandem mass spectrometry (HT-MS/MS) and inductively coupled plasma mass spectrometry (ICP-MS).”

This is a huge surprise. It reveals how naive we are about the wide range of chemistries that microbes do.

—John Tainer of Berkeley Lab’s Life Sciences Division and the Scripps Research Institute in La Jolla, CA

The team catalogued the metals in three microbes: Escherichia coli, Sulfolobus solfataricus (from a hotspring in Yellowstone National Park) and Pyrococcus furiosus (an extremophile that thrives in undersea thermal vents).

As an example, they found that of 343 metal peaks in chromatography fractions from P. furiosus, 158 did not match any predicted metalloprotein. Unassigned peaks included metals known to be used (cobalt, iron, nickel, tungsten and zinc; 83 peaks) plus metals the organism was not thought to assimilate (lead, manganese, molybdenum, uranium and vanadium; 75 peaks).

The scientists traced these newfound metals to the proteins that contain them, called metalloproteins. They discovered four new metalloproteins in the microbe, which increased the number of known metalloproteins in P. furiosus by almost a quarter. Their discovery also increased the number of nickel-containing enzymes in all of biology from eight to ten.

A similar survey of the other two microbes unearthed additional unexpected metals and new metalloproteins. Based on these findings, the team suggests that metalloproteins are much more extensive and diverse in the microbial world than scientists realized.

We thought we knew most of the metalloproteins out there. But it turns out we only know a tiny fraction of them. We now have to look at microbial genomes with a fresh eye.

—John Tainer

The team used a novel combination of two techniques in their process. Biochemical fractionation enabled them to take apart a microbe while keeping its proteins intact and stable, ready to be analyzed in their natural state. Next, a technology called inductively coupled plasma mass spectrometry allowed them to identify extremely low quantities of individual metals in these proteins. Together, these tools provide a quick tally of all of the metalloproteins in a microbe.

The current method for discovering metalloproteins is much slower: genetically sequencing a microbe, identifying the proteins encoded by its genes, and structurally characterizing each protein.

In addition to gaining a better understanding of the biochemical diversity of microbes, the team’s new metal-hunting technique could expedite the search for new biochemical capabilities in microbial life that can be harnessed for clean energy development, carbon sequestration, and other applications.

If you want to degrade cellulose to make biofuel, and you know the enzymes involved require a specific metal-driven chemistry, then you can use this technique to find those enzymes in microbes.

—Steven Yannone

The research was funded by the Department of Energy Office of Science.


  • Aleksandar Cvetkovi, et al. (2010) Microbial metalloproteomes are largely uncharacterized.Nature advance online publication doi: 10.1038/nature09265