{"id":3483,"date":"2019-05-09T15:12:50","date_gmt":"2019-05-09T06:12:50","guid":{"rendered":"http:\/\/163.180.4.222\/lab\/?p=3483"},"modified":"2019-05-09T15:12:50","modified_gmt":"2019-05-09T06:12:50","slug":"an-on-switch-for-proteins","status":"publish","type":"post","link":"https:\/\/biochemistry.khu.ac.kr\/lab\/?p=3483","title":{"rendered":"An \u2018on\u2019 switch for proteins"},"content":{"rendered":"<p>&nbsp;<\/p>\n<h5><\/h5>\n<h5>Current methods for producing proteins that can be activated by light require knowledge of the protein\u2019s active site, or can reduce the protein\u2019s functionality. A technique that overcomes these issues has been devised.<\/h5>\n<p>&nbsp;<\/p>\n<div class=\"article__body serif cleared\">\n<p>Cells can activate the same proteins at different times or places to generate diverse effects \u2014 for example, the same enzymes can be involved in both cell growth and programmed cell death. Many cellular processes that depend on the timing and site of protein activity can be studied in living cells, by triggering localized protein activity and examining the effects. In the past few years, scientists have developed \u2018photoactivation\u2019 methods that allow protein functions to be switched on by light<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR1\">1<\/a><\/sup><sup>,<\/sup><sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR2\">2<\/a><\/sup>.\u00a0<a href=\"https:\/\/www.nature.com\/articles\/s41586-019-1188-1\" data-track=\"click\" data-label=\"https:\/\/www.nature.com\/articles\/s41586-019-1188-1\" data-track-category=\"body text link\">Writing in\u00a0<i>Nature<\/i><\/a>, Wang\u00a0<i>et al<\/i>.<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR3\">3<\/a><\/sup>\u00a0now describe a photoactivation strategy that is both broadly applicable and minimally perturbs normal protein functions.<\/p>\n<p>&nbsp;<\/p>\n<aside class=\"recommended pull pull--left sans-serif\" data-label=\"Related\"><a href=\"https:\/\/www.nature.com\/articles\/s41586-019-1188-1\" data-track=\"click\" data-track-label=\"recommended article\"><img decoding=\"async\" class=\"recommended__image\" src=\"https:\/\/media.nature.com\/w400\/magazine-assets\/d41586-019-01394-1\/d41586-019-01394-1_16690746.jpg\" \/><\/a><\/p>\n<p class=\"recommended__title serif\">Read the paper: Time-resolved protein activation by proximal decaging in living systems<\/p>\n<\/aside>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<p>Approaches for the photoactivation and photoinhibition of proteins are available, but it is often difficult to apply these without modifying some of the proteins\u2019 activities. These methods work by manipulating amino-acid residues involved in the target protein\u2019s mechanism of action, within the active site. However, the structure and active sites of many proteins are poorly understood, preventing such methods from being applied to many important systems.<\/p>\n<p>Wang and colleagues report a method that they term computationally aided and genetically encoded proximal decaging (CAGE-prox). In CAGE-prox, a straightforward computational method is used to identify a position in a protein of interest at which the introduction of a bulky chemical group is likely to perturb the protein\u2019s interaction with its substrate. The amino-acid residue at that position is then replaced with a tyrosine residue that has been modified<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR4\">4<\/a><\/sup>\u00a0to carry a group that can be cleaved using light. Once installed, this bulky group blocks the protein\u2019s activity until light irradiation \u2018prunes\u2019 it back to the normal tyrosine structure (Fig. 1), whereupon activity is restored.<\/p>\n<p>&nbsp;<\/p>\n<figure class=\"figure\">\n<div class=\"embed intensity--high\">\n<div class=\"embed intensity--high\"><img decoding=\"async\" class=\"figure__image\" src=\"https:\/\/media.nature.com\/w800\/magazine-assets\/d41586-019-01394-1\/d41586-019-01394-1_16698236.jpg\" alt=\"\" data-src=\"\/\/media.nature.com\/w800\/magazine-assets\/d41586-019-01394-1\/d41586-019-01394-1_16698236.jpg\" \/><\/div>\n<\/div><figcaption>\n<p class=\"figure__caption sans-serif\"><span class=\"mr10\"><b>Figure 1 | A method for activating proteins using light.<\/b>\u00a0Wang\u00a0<i>et al<\/i>.<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR3\">3<\/a><\/sup>\u00a0report a technique that they call computationally aided and genetically encoded proximal decaging (CAGE-prox), which activates proteins in cells. In CAGE-prox, an amino-acid residue close to a protein\u2019s active site is replaced by a modified tyrosine residue. The modified residue carries a bulky group on its side chain, which prevents the protein\u2019s substrate (in this case, another protein; green) from binding in the active site. Light clips off the bulky side chain, leaving a normal tyrosine residue that allows substrate binding, thereby activating the protein.<\/span><\/p>\n<\/figcaption><\/figure>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<p>The computational method requires no information about a protein\u2019s mechanism of action. CAGE-prox can therefore be applied to an amazingly wide range of target proteins. The authors elected to use a modified tyrosine residue, rather than other amino acids that could be modified with a light-cleavable group, because they found that the normal tyrosine produced after cleavage proved least likely to perturb folding or normal protein binding.<\/p>\n<p>One of the most striking advantages of CAGE-prox is its ability to produce an almost-native protein analogue \u2014 irradiation produces a protein that differs from the normal one by only a single amino-acid residue, which need not be in the active site. Other approaches have used light to link a fragment of an active protein to a second protein anchored at a specific site in the cell, thereby driving the fragment to specific regions<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR1\">1<\/a><\/sup>. Alternatively, engineered protein domains that change shape when irradiated have been inserted into a target protein to alter its conformation when illuminated, or have been positioned so that they block the target\u2019s active site only in the dark<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR1\">1<\/a><\/sup><sup>,<\/sup><sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR2\">2<\/a><\/sup>. These previously reported methods alter the overall structure of the target protein, thus potentially affecting its ability to interact normally with its biological partners.<\/p>\n<p>By contrast, the surfaces used by wild-type proteins to mediate interactions with other cell components are retained almost intact in the CAGE-prox proteins after irradiation. This allows the modified proteins to target their normal binding partners in cells, and to be simultaneously activated at multiple locations in the same way as the wild-type protein.<\/p>\n<p>It is perhaps surprising that the small changes associated with just one light-sensitive tyrosine residue can affect the interactions of a target protein so effectively. The success of Wang and colleagues\u2019 method depends crucially on the ability to select the best site for modification. Encouragingly, the computational modelling used in CAGE-prox identified fewer than ten possible modification sites for each of the diverse proteins studied, limiting the number of amino-acid positions that had to be tested experimentally to find the most effective one.<\/p>\n<p>The researchers used CAGE-prox not only to activate diverse protein structures, but also to control the sensitivity of kinase enzymes to inhibitors, thereby allowing modified and wild-type kinases to be inhibited independently. Just as impressively, Wang\u00a0<i>et al<\/i>. used their method to activate proteins that have anticancer activity, and showed that light-triggered activation of these proteins inhibits tumour growth\u00a0<i>in vivo<\/i>\u00a0in mice.<\/p>\n<p>In other photoactivation methods, a key amino-acid residue in the active site is identified and replaced by a modified version of that residue. By contrast, in CAGE-prox, the identified residue does not have to be in the active site and is always replaced by the same modified tyrosine residue. This tyrosine is introduced using a cell-based technique called unnatural-amino-acid mutagenesis<sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR5\">5<\/a><\/sup><sup>,<\/sup><sup><a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29#ref-CR6\">6<\/a><\/sup>. The use of this technique could be seen as a weakness of Wang and co-workers\u2019 approach, because unnatural-amino-acid mutagenesis is not suitable for all cell types, and can require substantial optimization for each application. Furthermore, the covalent bond that is cleaved to remove the bulky side chain from the light-sensitive tyrosine residue can be broken only by using high-energy light (wavelengths of less than 400 nanometres), which is toxic to living cells. These are likely to be short-term obstacles, however, because many laboratories are actively pursuing and improving methods for altering proteins in cultured cells, and even\u00a0<i>in vivo<\/i>.<\/p>\n<p>With its remarkable simplicity and generality, CAGE-prox opens the door to studies of previously inaccessible cellular pathways, and of the spatio-temporal control of processes that determine cell behaviour. The range of applications that Wang\u00a0<i>et al.<\/i>\u00a0have already proved in principle for their technique is remarkable. No doubt, many more will soon follow.<\/p>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<\/div>\n<div class=\"emphasis\">doi: 10.1038\/d41586-019-01394-1<\/div>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<p>(\uc6d0\ubb38: <a href=\"https:\/\/www.nature.com\/articles\/d41586-019-01394-1?utm_source=feedburner&amp;utm_medium=feed&amp;utm_campaign=Feed%3A+nature%2Frss%2Fcurrent+%28Nature+-+Issue%29\">\uc5ec\uae30<\/a>\ub97c \ud074\ub9ad\ud558\uc138\uc694~)<\/p>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n<p>&nbsp;<\/p>\n","protected":false},"excerpt":{"rendered":"<p>&nbsp; Current methods for producing proteins that can be activated by light require knowledge of the protein\u2019s active site, or can reduce the protein\u2019s functionality.<a href=\"https:\/\/biochemistry.khu.ac.kr\/lab\/?p=3483\" class=\"more-link\">(more&#8230;)<\/a><\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"_monsterinsights_skip_tracking":false,"_monsterinsights_sitenote_active":false,"_monsterinsights_sitenote_note":"","_monsterinsights_sitenote_category":0,"jetpack_post_was_ever_published":false,"_jetpack_newsletter_access":"","_jetpack_dont_email_post_to_subs":false,"_jetpack_newsletter_tier_id":0,"_jetpack_memberships_contains_paywalled_content":false,"_jetpack_memberships_contains_paid_content":false,"footnotes":"","jetpack_publicize_message":"","jetpack_publicize_feature_enabled":true,"jetpack_social_post_already_shared":true,"jetpack_social_options":{"image_generator_settings":{"template":"highway","default_image_id":0,"font":"","enabled":false},"version":2}},"categories":[33,34,29,30],"tags":[],"class_list":["post-3483","post","type-post","status-publish","format-standard","hentry","category-do-biology","category-lets-do-chemistry","category-lets-do-science","category-recent-science-news"],"aioseo_notices":[],"jetpack_publicize_connections":[],"jetpack_featured_media_url":"","jetpack-related-posts":[{"id":1859,"url":"https:\/\/biochemistry.khu.ac.kr\/lab\/?p=1859","url_meta":{"origin":3483,"position":0},"title":"Programmable protein circuits in living cells","author":"biochemistry","date":"September 25, 2018","format":false,"excerpt":"\u00a0 \u00a0 Science\u00a0\u00a021 Sep 2018: Vol. 361, Issue 6408, pp. 1252-1258 DOI: 10.1126\/science.aat5062 \u00a0 \uc5ec\uae30\ub97c \ud074\ub9ad\ud558\uc138\uc694~ \u00a0 \u00a0 Building smarter synthetic biological circuits Synthetic genetic and biological regulatory circuits can enable logic functions to form the basis of biological computing; synthetic biology can also be used to control cell behaviors\u2026","rel":"","context":"In &quot;Let's Do Biology!&quot;","block_context":{"text":"Let's Do Biology!","link":"https:\/\/biochemistry.khu.ac.kr\/lab\/?cat=33"},"img":{"alt_text":"","src":"","width":0,"height":0},"classes":[]},{"id":2531,"url":"https:\/\/biochemistry.khu.ac.kr\/lab\/?p=2531","url_meta":{"origin":3483,"position":1},"title":"CRISPR adapted to respond to infected cells","author":"biochemistry","date":"January 18, 2019","format":false,"excerpt":"\u00a0 \u00a0 By making a small change to the sequence of the Cas9 protein researchers can control the enzyme\u2019s activity. Credit: B. L. Oakes\u00a0et al.\/Cell \u00a0 \u00a0 \u00a0 Engineered tweaks to the popular gene-editing system allow it to fight viral infection. \u00a0 A bacterial enzyme that researchers often use to\u2026","rel":"","context":"In &quot;Let's Do Biology!&quot;","block_context":{"text":"Let's Do Biology!","link":"https:\/\/biochemistry.khu.ac.kr\/lab\/?cat=33"},"img":{"alt_text":"","src":"","width":0,"height":0},"classes":[]},{"id":2250,"url":"https:\/\/biochemistry.khu.ac.kr\/lab\/?p=2250","url_meta":{"origin":3483,"position":2},"title":"Artificial cells gain communication skills","author":"biochemistry","date":"December 3, 2018","format":false,"excerpt":"\u00a0 \u00a0 No biologist would mistake the microscopic \u201ccells\u201d that chemical biologist Neal Devaraj and colleagues are whipping up at the University of California, San Diego (UCSD), for the real thing. 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