{"id":125350,"date":"2023-04-17T07:12:00","date_gmt":"2023-04-17T05:12:00","guid":{"rendered":"https:\/\/renewable-carbon.eu\/news\/?p=125350"},"modified":"2023-04-13T09:12:06","modified_gmt":"2023-04-13T07:12:06","slug":"structure-of-oil-eating-enzyme-opens-door-to-bioengineered-catalysts","status":"publish","type":"post","link":"https:\/\/renewable-carbon.eu\/news\/structure-of-oil-eating-enzyme-opens-door-to-bioengineered-catalysts\/","title":{"rendered":"Structure of ‘Oil-Eating’ Enzyme Opens Door to Bioengineered Catalysts"},"content":{"rendered":"\n\n\n

Scientists at the U.S. Department of Energy\u2019s Brookhaven National Laboratory have produced the first atomic-level structure of an enzyme that selectively cuts carbon-hydrogen bonds\u2014the first and most challenging step in turning simple hydrocarbons into more useful chemicals. As described in a paper just published in\u00a0Nature Structural & Molecular Biology<\/em>, the detailed atomic level \u201cblueprint\u201d suggests ways to engineer the enzyme to produce desired products.<\/p>\n\n\n\n

\"\"
Long-sought structure of oil-eating enzyme complex:<\/strong>\u00a0A high-resolution cryo-EM map of the transmembrane two-protein complex (left) allows researchers to determine the locations of individual amino acids that make up the two proteins (right). AlkG (gray) serves and an electron carrier, transporting electrons from its single iron atom (red sphere) to the two iron atoms (red spheres) at the active site of the AlkB enzyme (colorful ribbon structure). The magenta structure below the active site is the substrate (see close-up views). \u00a9<\/strong> Brookhaven National Laboratory<\/figcaption><\/figure>\n\n\n\n

\u201cWe want to create a diverse pool of biocatalysts where you can specifically select the desired substrate to produce wanted and unique products from abundant hydrocarbons,\u201d said study co-lead Qun Liu, a Brookhaven Lab structural biologist. \u201cThe approach would give us a controllable way to convert cheap and abundant alkanes\u2014simple carbon-hydrogen compounds that make up 20-50 percent of crude oil\u2014into more valuable bioproducts or chemical precursors, including alcohols, aldehydes, carboxylates, and epoxides.\u201d<\/p><\/blockquote>\n\n\n\n

The idea is particularly attractive because most industrial catalytic processes used for alkane conversions produce unwanted byproducts and heat-trapping carbon dioxide (CO2<\/sub>) gas. They also contain costly materials and require high temperatures and pressure. The biological enzyme, known as AlkB, operates under more ordinary conditions and with very high specificity. It uses inexpensive earth-abundant iron to initiate the chemistry while producing few unwanted byproducts.<\/p>\n\n\n\n

\u201cNature has figured out how to do this kind of chemistry with an inexpensive abundant metal and at ambient temperature and pressures,\u201d said John Shanklin, chair of Brookhaven Lab\u2019s Biology Department and a senior author on the paper. \u201cAs a result, there\u2019s been massive interest in this enzyme, but a complete lack of understanding of its architecture and how it works\u2014which is necessary to re-engineer it for new purposes. With this structure, we have now overcome this obstacle.\u201d<\/p><\/blockquote>\n\n\n\n

From rancid oil to sweet success<\/h3>\n\n\n\n
\"\"
Research team:<\/strong>\u00a0Brookhaven Lab scientists Jin Chai, Qun Liu, John Shanklin, and Sean McSweeney stand in front of the cryo-electron microscope (cryo-EM) used to decipher the long-sought structure of an enzyme that selectively cleaves hydrocarbon bonds. \u00a9<\/strong> Brookhaven National Laboratory<\/figcaption><\/figure><\/div>\n\n\n\n

AlkB was discovered 50 years ago in a machine shop, where bacteria were digesting cooling oil making it smell rancid. Biochemists discovered the bacterial enzyme AlkB as the factor enabling the microbes\u2019 unusual appetite. Scientists have been interested in harnessing AlkB\u2019s hydrocarbon-chomping ability ever since.<\/p>\n\n\n\n

Over the years, studies revealed that the enzyme sits partially embedded in the bacteria\u2019s membranes, and that it operates in conjunction with two other proteins. Shanklin and Liu\u2014and scientists elsewhere\u2014tried solving the enzyme\u2019s structure using x-ray crystallography. That method bounces high-intensity x-rays off a crystallized version of a protein to identify where the atoms are. But membrane proteins like AlkB are notoriously difficult to crystallize\u2014especially when they are part of a multi-protein complex.<\/p>\n\n\n\n

\u201cWe couldn\u2019t get high enough resolution,\u201d Liu said.<\/p>\n\n\n\n

Then in early 2021, Brookhaven opened its new cryo-electron microscope (cryo-EM) facility, the Laboratory for BioMolecular Structure<\/a> (LBMS). The scientists used a cryo-EM, which does not require a crystallized sample, to take pictures of a few million individual frozen protein molecules from many different angles. Computational tools then sorted through the images, identified and averaged the common features\u2014and ultimately generated a high-resolution, three-dimensional map of the enzyme complex. Using this map, the scientists then pieced together the known atomic-level structures of the individual amino acids that make up the protein complex to fill in the details in three dimensions.<\/p>\n\n\n\n

Identifying the right conditions to stabilize the transmembrane region of the enzyme and maintain the structural details was a challenge that required a good deal of trial and error. Shanklin credits Jin Chai, one of the researchers in his lab, \u201cfor his commitment and determination to solving this puzzle.\u201d   <\/p>\n\n\n\n

Structure reveals how enzyme works<\/h3>\n\n\n\n

The detailed structure shows exactly how AlkB and one of the two associated proteins (AlkG) work together to cleave carbon-hydrogen bonds. In fact, the solved structure contained an unexpected bonus: a substrate alkane molecule that was trapped in the enzyme\u2019s active site cavity.  <\/p>\n\n\n\n

\"\"
Active site:<\/strong>\u00a0These closeups of the AlkB active site show how nine histidine amino acids (denoted as “H” in the left image) form a cavity (gray shaded region, right). This cavity guides the substrate (magenta) to the active site (near the two iron, Fe, atoms) in a single orientation, where only the terminal carbon-hydrogen bond can be cleaved. Modifying the enzyme to change the shape of this cavity could allow the enzyme to attack different C-H bonds. \u00a9<\/strong> Brookhaven National Laboratory<\/figcaption><\/figure><\/div>\n\n\n\n

\u201cOur structure shows how the amino acids that make up this enzyme form a cavity that orients the hydrocarbon substrate so that just one specific carbon-hydrogen bond can approach the active site,\u201d Liu said. \u201cIt also shows how electrons move from the carrier protein (AlkG) to the di-iron center at the enzyme\u2019s active site, allowing it to activate a molecule of oxygen to attack this bond.\u201d<\/p><\/blockquote>\n\n\n\n

Shanklin suggests thinking of the enzyme as a bond-cutting machine like a circular saw: \u201cHow you hold the alkane with respect to the enzyme\u2019s di-iron center determines how the activated oxygen interacts with the hydrocarbon. If you guide the end of the alkane against the activated oxygen, it\u2019s going to initiate some chemistry on that last carbon.<\/p>\n\n\n\n

\u201cThe engineering we want to do is to change the shape of the active site cavity so we can have the substrate (or a different substrate) approach the activated oxygen at different angles and in different C-H bond locations to perform different reactions.\u201d<\/p><\/blockquote>\n\n\n\n

In nature, the scientists noted, a third protein not included in this structure (AlkT) provides the electrons to AlkG, the carrier protein. The carrier protein then transports the electrons to the two iron atoms that activate oxygen at AlkB\u2019s active site. Replacing that electron donating protein with an electrode to supply electrons would be simpler and less costly than using the biological electron donor, they suggest.<\/p>\n\n\n\n

DOE just funded the team\u2019s proposal to develop such \u2018Transformative Biohybrid Diiron Catalysts for C-H Bond Functionalization,\u2019<\/a> based in part on this preliminary structural work.<\/p>\n\n\n\n

\u201cThis structure and our knowledge of how the AlkG\/AlkB complex works, puts us in a great position to bioengineer this enzyme to select which carbon-hydrogen bond gets activated in a variety of substrates and to control the electrons and oxygen to re-engineer its selectivity,\u201d Liu said. \u00a0\u00a0\u00a0<\/p><\/blockquote>\n\n\n\n

This work was supported by the DOE Office of Science (BES) and by Laboratory Directed Research and Development funds at Brookhaven Lab. LBMS is supported by the DOE Office of Science (BER). This research also used resources of Brookhaven Lab\u2019s Center for Functional Nanomaterials (CFN), which is a U.S. Department of Energy Office of Science (BES) User Facility.<\/p>\n\n\n\n

Brookhaven National Laboratory is supported by the Office of Science of the U.S. Department of Energy. The Office of Science\u00a0is the single largest supporter of basic research in the physical sciences in the United States and is working to address some of the most pressing challenges of our time. <\/em><\/p>\n","protected":false},"excerpt":{"rendered":"

Scientists at the U.S. Department of Energy\u2019s Brookhaven National Laboratory have produced the first atomic-level structure of an enzyme that selectively cuts carbon-hydrogen bonds\u2014the first and most challenging step in turning simple hydrocarbons into more useful chemicals. As described in a paper just published in\u00a0Nature Structural & Molecular Biology, the detailed atomic level \u201cblueprint\u201d suggests […]<\/p>\n","protected":false},"author":3,"featured_media":0,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"_seopress_robots_primary_cat":"none","nova_meta_subtitle":"Atomic level details reveal how enzyme selectively breaks hydrocarbon bonds, suggesting bioengineering strategies for making useful chemicals","footnotes":""},"categories":[5572],"tags":[12753,5796,15152,5840,12569],"supplier":[4469,4909,22023,22022,11236,4116],"class_list":["post-125350","post","type-post","status-publish","format-standard","hentry","category-bio-based","tag-bioengineering","tag-biotechnology","tag-catalyst","tag-enzymes","tag-oil","supplier-brookhaven-national-laboratory","supplier-center-for-functional-nanomaterials-cfn","supplier-laboratory-for-biomolecular-structure-lbms","supplier-nature-structural-molecular-biology","supplier-u-s-department-of-energy","supplier-us-doe-office-of-science-sc"],"_links":{"self":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts\/125350","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/users\/3"}],"replies":[{"embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/comments?post=125350"}],"version-history":[{"count":0,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts\/125350\/revisions"}],"wp:attachment":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/media?parent=125350"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/categories?post=125350"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/tags?post=125350"},{"taxonomy":"supplier","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/supplier?post=125350"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}