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<input type="email" aria-label="Sender Email Address" placeholder="email@example.com" id="email-from" class="email-from" pattern="^[a-zA-Z0-9_.+-]+@[a-zA-Z0-9-]+\.[a-zA-Z0-9-.]+$" maxlength="256">
</div>
<div class="action-panel-control-wrap">
<label for="email-citation-format" class="action-panel-label">
Format:
</label>
<select id="email-citation-format" name="citation-format" class="action-panel-selector email-citation-format">
<option selected="selected" value="summary">Summary</option>
<option value="summary-text">Summary (text)</option>
<option value="abstract">Abstract</option>
<option value="abstract-text">Abstract (text)</option>
</select>
</div>
<div class="include-supplemental-container">
<input type="checkbox" aria-label="Include MeSH and other data" name="include-supplemental" id="email-include-supplemental" class="email-include-supplemental">
<label for="email-include-supplemental" class="email-include-supplemental-label">MeSH and other data</label>
</div>
<div class="form-field recaptcha ">
<div class="g-recaptcha" id="id-recaptcha" data-sitekey="6LfsWHMdAAAAAClKbtOpjQ2pMjgsGxvv7NdZW9uI"></div>
</div>
<div id="captcha-error-message" class="usa-input-error-message captcha-validation-message" role="alert"></div>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Sending..."
data-ga-category="save_share"
data-ga-action="email"
data-ga-label="send">
Send email
</button>
<button class="action-panel-cancel"
aria-label="Close 'Email citations' panel"
ref="linksrc=close_email_panel"
aria-controls="email-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="email"
data-ga-label="cancel">
Cancel
</button>
</div>
<input type="hidden" name="email-search-details" value="" />
<input type="hidden" name="email-search-details-hash" value="0e42663a6c3bd85498fcb88798998fed7bfdc45d457db35281e41afe13cc0524" />
</form>
</div>
</div>
<div id="collections-action-panel"
class="collections-action-panel action-panel in-progress-dots-panel"
aria-hidden="true"
data-collections-open-panel-enabled="false"
data-collections-open-panel-url-hash="#open-collections-panel">
<div class="inner-wrap">
<h3 class="action-panel-heading">
Add to Collections
</h3>
<form id="collections-action-panel-form"
class="collections-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
data-existing-collections-url="/list-existing-collections/"
data-add-to-existing-collection-url="/add-to-existing-collection/"
data-create-and-add-to-new-collection-url="/create-and-add-to-new-collection/"
data-get-article-ids-by-search-url="/get-article-ids-by-search/"
data-myncbi-max-collection-name-length="100"
data-add-to-collection-max-amount="1000"
data-collections-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/">
<input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI">
<div class="choice-group" role="radiogroup">
<ul class="radio-group-items">
<li>
<input type="radio"
id="collections-action-panel-new"
class="collections-new"
name="collections"
value="new"
data-ga-category="save_share"
data-ga-action="collections"
data-ga-label="collections_radio_new">
<label for="collections-action-panel-new">Create a new collection</label>
</li>
<li>
<input type="radio"
id="collections-action-panel-existing"
class="collections-existing"
name="collections"
value="existing"
checked="true"
data-ga-category="save_share"
data-ga-action="collections"
data-ga-label="collections_radio_existing">
<label for="collections-action-panel-existing">Add to an existing collection</label>
</li>
</ul>
</div>
<div class="controls-wrapper">
<div class="action-panel-control-wrap new-collections-controls">
<label for="collections-action-panel-add-to-new" class="action-panel-label required-field-asterisk">
Name your collection:
</label>
<input
type="text"
name="add-to-new-collection"
id="collections-action-panel-add-to-new"
class="collections-action-add-to-new"
pattern="[^&quot;&amp;=&lt;&gt;\/]*" title="The following characters are not allowed in the Name field: &quot;&amp;=&lt;&gt;/"
maxlength="100"
data-ga-category="save_share"
data-ga-action="create_collection"
data-ga-label="non_favorties_collection">
<div class="collections-new-name-too-long usa-input-error-message selection-validation-message">
Name must be less than 100 characters
</div>
</div>
<div class="action-panel-control-wrap existing-collections-controls">
<label for="collections-action-panel-add-to-existing" class="action-panel-label">
Choose a collection:
</label>
<select id="collections-action-panel-add-to-existing"
class="action-panel-selector collections-action-add-to-existing"
data-ga-category="save_share"
data-ga-action="select_collection"
data-ga-label="($('#collections-action-add-to-existing').val() === 'Favorites') ? 'Favorites' : 'non_favorites_collection'">
</select>
<div class="collections-retry-load-on-error usa-input-error-message selection-validation-message">
Unable to load your collection due to an error<br>
<a href="#">Please try again</a>
</div>
</div>
</div>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Adding..."
data-pinger-ignore
data-ga-category="save_share"
data-ga-action="collections"
data-ga-label="add">
Add
</button>
<button class="action-panel-cancel"
aria-label="Close 'Add to Collections' panel"
ref="linksrc=close_collections_panel"
aria-controls="collections-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="collections"
data-ga-label="cancel">
Cancel
</button>
</div>
</form>
</div>
</div>
<div id="bibliography-action-panel"
class="bibliography-action-panel action-panel in-progress-dots-panel"
aria-hidden="true"
data-bibliography-open-panel-enabled="false"
data-bibliography-open-panel-url-hash="#open-bibliography-panel">
<div class="inner-wrap">
<h3 class="action-panel-heading">
Add to My Bibliography
</h3>
<form id="bibliography-action-panel-form"
class="bibliography-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
data-add-to-bibliography-max-amount="100"
data-add-to-bibliography-batch-size="10"
data-bibliography-delegates-url="/list-bibliography-delegates/"
data-add-to-bibliography-url="/add-to-bibliography/"
data-get-article-ids-by-search-url="/get-article-ids-by-search/"
data-mybib-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/mybibliography/">
<input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI">
<div class="action-panel-control-wrap bibliographies-controls">
<div class="choice-group">
<ul class="bibliographies-action-add radio-group-items">
<li>
<input name="bibliography" id="my-bibliography" class="my-bibliography" type="radio" checked/>
<label for="my-bibliography">My Bibliography</label>
</li>
</ul>
</div>
</div>
<div class="bibliographies-retry-load-on-error usa-input-error-message selection-validation-message">
Unable to load your delegates due to an error<br>
<a href="#">Please try again</a>
</div>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Adding..."
data-pinger-ignore>
Add
</button>
<button class="action-panel-cancel"
aria-label="Close 'Add to bibliography' panel"
ref="linksrc=close_bibliography_panel"
aria-controls="bibliography-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="mybib"
data-ga-label="cancel">
Cancel
</button>
</div>
</form>
</div>
</div>
<div id="saved-search-action-panel" class="saved-search-action-panel action-panel " aria-hidden="true"
data-saved-search-open-panel-enabled="false"
data-saved-search-open-panel-url-hash="#open-saved-search-panel">
<div class="inner-wrap">
<h2 class="action-panel-heading">
Your saved search
</h2>
<form id="saved-search-action-panel-form"
class="saved-search-action-panel-form action-panel-content action-form"
data-create-saved-search-url="/create-saved-search/"
data-try-search-terms-url="/try-search-term/"
data-saved-search-root-url="https://www.ncbi.nlm.nih.gov/myncbi/searches/">
<input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI">
<div class="action-panel-control-wrap">
<label for="saved-search-name" class="action-panel-label saved-search-name-label required-field-asterisk">
Name of saved search:
</label>
<input maxlength="200"
type="text"
name="saved-search-name"
id="saved-search-name"
class="saved-search-name"
value=""
required
pattern="[^&quot;&amp;=&lt;&gt;\/]*" title="The following characters are not allowed in the Name field: &quot;&amp;=&lt;&gt;/">
</div>
<div class="action-panel-control-wrap">
<label for="saved-search-term" class="action-panel-label required-field-asterisk">
Search terms:
</label>
<textarea name="saved-search-term" id="saved-search-term" class="saved-search-term" required></textarea>
</div>
<div class="test-search-term-wrap">
<a href="#" class="try-search-term">Test search terms</a>
</div>
<div class="choice-group action-panel-extra-margin-top">
<span class="action-panel-label" id="fieldset-label">
Would you like email updates of new search results?
</span>
<fieldset id="saved-search-alert" aria-describedby="fieldset-label">
<legend class="usa-sr-only">Saved Search Alert Radio Buttons</legend>
<ul class="radio-group-items">
<li>
<input type="radio" id="saved-search-alert-yes" class="saved-search-alert-yes" name="saved-search-alert" value="yes" checked>
<label for="saved-search-alert-yes" class="action-panel-label">Yes</label>
</li>
<li>
<input aria-label="No radio input" type="radio" id="saved-search-alert-no" class="saved-search-alert-no" name="saved-search-alert" value="no">
<label for="saved-search-alert-no" class="action-panel-label">No</label>
</li>
</ul>
</fieldset>
</div>
<div class="alert-schedule-wrap">
<div class="action-panel-control-wrap">
<label class="action-panel-label">
Email:
</label>
<span aria-label="Email address" id="saved-search-email" class="action-panel-label"><span class="action-panel-label-bold"></span> (<a class="myncbi-account-settings" href="https://www.ncbi.nlm.nih.gov/account/settings/">change</a>)</span>
</div>
<div class="action-panel-control-wrap action-panel-extra-margin-top">
<label for="saved-search-frequency" class="action-panel-label">
Frequency:
</label>
<select id="saved-search-frequency" class="no-border-panel-selector saved-search-frequency">
<option value="monthly">Monthly</option>
<option value="weekly">Weekly</option>
<option value="daily">Daily</option>
</select>
</div>
<div class="action-panel-control-wrap saved-search-monthly-additional">
<label for="saved-search-monthly-on-day" class="action-panel-label">
Which day?
</label>
<select id="saved-search-monthly-on-day" class="no-border-panel-selector">
<option value="Sunday">The first Sunday</option>
<option value="Monday">The first Monday</option>
<option value="Tuesday">The first Tuesday</option>
<option value="Wednesday">The first Wednesday</option>
<option value="Thursday">The first Thursday</option>
<option value="Friday">The first Friday</option>
<option value="Saturday">The first Saturday</option>
<option value="day">The first day</option>
<option value="weekday">The first weekday</option>
</select>
</div>
<div class="action-panel-control-wrap saved-search-weekly-additional">
<label for="saved-search-weekly-on-day" class="action-panel-label">
Which day?
</label>
<select id="saved-search-weekly-on-day" class="no-border-panel-selector saved-search-weekly-on-day">
<option value="Sunday">Sunday</option>
<option value="Monday">Monday</option>
<option value="Tuesday">Tuesday</option>
<option value="Wednesday">Wednesday</option>
<option value="Thursday">Thursday</option>
<option value="Friday">Friday</option>
<option value="Saturday">Saturday</option>
</select>
</div>
<div class="action-panel-control-wrap">
<label for="saved-search-report" class="action-panel-label">
Report format:
</label>
<select id="saved-search-report" class="no-border-panel-selector saved-search-report">
<option value="DocSum">Summary</option>
<option value="DocSumText">Summary (text)</option>
<option value="Abstract">Abstract</option>
<option value="AbstractText">Abstract (text)</option>
<option value="MEDLINE">PubMed</option>
</select>
</div>
<div class="action-panel-control-wrap">
<label for="saved-search-amount" class="action-panel-label">
Send at most:
</label>
<select id="saved-search-amount" class="no-border-panel-selector saved-search-amount">
<option value="1">1 item</option>
<option value="5" selected>5 items</option>
<option value="10">10 items</option>
<option value="20">20 items</option>
<option value="50">50 items</option>
<option value="100">100 items</option>
<option value="200">200 items</option>
</select>
</div>
<div>
<input type="checkbox" id="saved-search-send-if-no-result" class="saved-search-send-if-no-result" name="saved-search-send-if-no-result">
<label for="saved-search-send-if-no-result" class="action-panel-label smaller-checkbox">
Send even when there aren't any new results
</label>
</div>
<div class="action-panel-control-wrap option-text-in-email-wrap">
<label for="saved-search-email-text" class="action-panel-label">
Optional text in email:
</label>
<textarea name="saved-search-email-text"
id="saved-search-email-text"
class="saved-search-email-text"></textarea>
</div>
</div>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Saving..."
data-ga-category="save_share"
data-ga-action="alert"
data-ga-label="save">
Save
</button>
<button class="action-panel-cancel"
aria-label="Close 'Your saved search' panel"
ref="linksrc=close_saved_search_panel"
aria-controls="saved-search-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="alert"
data-ga-label="cancel">
Cancel
</button>
</div>
</form>
</div>
</div>
<div id="citation-manager-action-panel" class="citation-manager-action-panel action-panel" aria-hidden="true">
<div class="inner-wrap">
<h2 class="action-panel-heading">
Create a file for external citation management software
</h2>
<form id="citation-manager-action-panel-form"
class="action-panel-content action-form"
action="/results-export-ids/"
data-by-search-action="/results-export-search-data/"
data-by-ids-action="/results-export-ids/"
method="post"
data-by-search-method="post"
data-by-ids-method="post">
<input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI">
<input name="results-format" type="hidden" value="pubmed"/>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Sending..."
data-ga-category="save_share"
data-ga-action="citation_manager"
data-ga-label="save">
Create file
</button>
<button class="action-panel-cancel"
aria-label="Close 'Send citations to citation manager' panel"
ref="linksrc=close_citation_manager_panel"
aria-controls="citation-manager-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="citation_manager"
data-ga-label="cancel">
Cancel
</button>
</div>
</form>
</div>
</div>
<div id="rss-action-panel" class="rss-action-panel action-panel " aria-hidden="true">
<div class="inner-wrap">
<h2 class="action-panel-heading">
Your RSS Feed
</h2>
<form id="rss-action-panel-form"
class="rss-action-panel-form action-panel-content action-form"
data-create-rss-feed-url="/create-rss-feed-url/"
data-search-form-term-value="">
<input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI">
<div class="action-panel-control-wrap">
<label for="rss-name" class="action-panel-label required-field-asterisk">
Name of RSS Feed:
</label>
<input maxlength="200"
placeholder="Name"
type="text"
name="rss-name"
id="rss-name"
class="rss-name"
value=''
required
pattern="[^&quot;&amp;=&lt;&gt;\/]*" title="The following characters are not allowed in the Name field: &quot;&amp;=&lt;&gt;/">
</div>
<div class="rss-limit-wrap">
<div class="action-panel-control-wrap action-panel-extra-margin-top">
<label for="rss-limit" class="action-panel-label">
Number of items displayed:
</label>
<select id="rss-limit" class="no-border-panel-selector rss-limit">
<option value="5">5</option>
<option value="10">10</option>
<option value="15" selected="selected">15</option>
<option value="20">20</option>
<option value="50">50</option>
<option value="100">100</option>
</select>
</div>
</div>
<div class="action-panel-actions">
<button class="action-panel-submit"
type="submit"
data-loading-label="Creating..."
data-ga-category="save_share"
data-ga-action="alert"
data-ga-label="save">
Create RSS
</button>
<button class="action-panel-cancel"
aria-label="Close 'Your RSS' panel"
ref="linksrc=close_rss_panel"
aria-controls="rss-action-panel"
aria-expanded="false"
data-ga-category="save_share"
data-ga-action="alert"
data-ga-label="cancel">
Cancel
</button>
</div>
<div class="action-panel-control-wrap rss-link-copy-wrap">
<label for="rss-link" class="usa-sr-only">RSS Link</label>
<input placeholder="Your RSS Feed Link" type="text" name="rss-link" id="rss-link" class="rss-link" title="RSS Link">
<button
type="button"
disabled
class="rss-link-copy-button disabled"
data-ga-category="save_share"
data-ga-action="rss"
data-ga-label="copy">
Copy
</button>
</div>
</form>
</div>
</div>
</div>
</div>
<div class="article-page" id="article-page" data-article-pmid="11078524">
<aside class="page-sidebar">
<div class="inner-wrap">
<div class="full-text-links">
<div class="full-view">
<h3 class="title">
Full text links
</h3>
<div class="full-text-links-list">
<a class="link-item
dialog-focus"
href="https://www.pnas.org/doi/10.1073/pnas.240347797?url_ver=Z39.88-2003&amp;rfr_id=ori:rid:crossref.org&amp;rfr_dat=cr_pub 0pubmed"
target="_blank"
rel="noopener"
ref="linksrc=fulltextorjournal_fulltext&amp;is_pmc=False&amp;PrId=4487&amp;itool=Abstract-def&amp;log$=linkouticon&amp;uid=11078524&amp;db=pubmed&amp;nlmid=7505876"
title="See full text options at Atypon"
data-ga-category="full_text"
data-ga-action="Atypon"
data-ga-label="11078524"
><img src="https://cdn.ncbi.nlm.nih.gov/corehtml/query/egifs/https:--pnas.org-pb-assets-images-pubmed-pnas-pubmed-fulltext.png" alt="Atypon full text link"><span class="text">
Atypon
</span></a><a class="link-item
pmc
"
href="https://pmc.ncbi.nlm.nih.gov/articles/pmid/11078524/"
target="_blank"
rel="noopener"
ref="linksrc=fulltextorjournal_fulltext&amp;is_pmc=True&amp;PrId=3494&amp;itool=Abstract-def&amp;log$=linkouticon&amp;uid=11078524&amp;db=pubmed&amp;nlmid=7505876"
title="Free full text at PubMed Central"
data-ga-category="full_text"
data-ga-action="PMC"
data-ga-label="11078524"
><span class="text">
Free PMC article
</span></a>
</div>
</div>
<div class="short-view">
<a href="#" class="full-text-links-button full-text-links-dialog-trigger">
Full text links
</a>
</div>
</div>
<div class="actions-buttons sidebar"><h3 class="title">Actions</h3><div class="inner-wrap"><button class="citation-button citation-dialog-trigger"
aria-label="Open dialog with citation text in different styles"
data-ga-category="save_share"
data-ga-action="cite"
data-ga-label="open"
data-all-citations-url="/11078524/citations/"
data-citation-style="nlm"
data-pubmed-format-link="/11078524/export/"><span class="button-label">Cite</span></button><link type="text/css" href="ncbi-overlay-block/src/overlay-block.css"><div class="collections-button-container" data-article-id="11078524" data-article-db="pubmed"><button class="collections-button collections-dialog-trigger"
aria-label="Save article in MyNCBI collections."
data-ga-category="collections_button"
data-ga-action="click"
data-ga-label="collections_button"
data-collections-open-dialog-enabled="false"
data-collections-open-dialog-url="https://account.ncbi.nlm.nih.gov/?back_url=https%3A%2F%2Fpubmed.ncbi.nlm.nih.gov%2F11078524%2F%23open-collections-dialog"
data-in-collections="false"><span class="button-label">Collections</span></button><div class="overlay" role="dialog"><div id="collections-action-dialog"
class="dialog collections-dialog"
aria-hidden="true"><div class="title">Add to Collections</div><div class="collections-action-panel action-panel"><form id="collections-action-dialog-form"
class="collections-action-panel-form action-panel-content action-form action-panel-smaller-selectors"
data-existing-collections-url="/list-existing-collections/"
data-add-to-existing-collection-url="/add-to-existing-collection/"
data-create-and-add-to-new-collection-url="/create-and-add-to-new-collection/"
data-myncbi-max-collection-name-length="100"
data-collections-root-url="https://www.ncbi.nlm.nih.gov/myncbi/collections/"><input type="hidden" name="csrfmiddlewaretoken" value="DNNSsUGKQ7ulx6NpX57YOLTYy3OcWtMoUAxX5K96qIhPrxTfAoK0iTpC67woP4kI"><div class="choice-group" role="radiogroup"><ul class="radio-group-items"><li><input type="radio"
id="collections-action-dialog-new"
class="collections-new"
name="collections"
value="new"
data-ga-category="collections_button"
data-ga-action="click"
data-ga-label="collections_radio_new"><label for="collections-action-dialog-new">Create a new collection</label></li><li><input type="radio"
id="collections-action-dialog-existing"
class="collections-existing"
name="collections"
value="existing"
checked="true"
data-ga-category="collections_button"
data-ga-action="click"
data-ga-label="collections_radio_existing"><label for="collections-action-dialog-existing">Add to an existing collection</label></li></ul></div><div class="controls-wrapper"><div class="action-panel-control-wrap new-collections-controls"><label for="collections-action-dialog-add-to-new" class="action-panel-label required-field-asterisk">
Name your collection:
</label><input
type="text"
name="add-to-new-collection"
id="collections-action-dialog-add-to-new"
class="collections-action-add-to-new"
pattern="[^&quot;&amp;=&lt;&gt;\/]*" title="The following characters are not allowed in the Name field: &quot;&amp;=&lt;&gt;/"
maxlength="100"
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<p>
Parkinson's disease is a common neurodegenerative disorder in which familial-linked genes have provided novel insights into the pathogenesis of this disorder. Mutations in Parkin, a ring-finger-containing protein of unknown function, are implicated in the pathogenesis of autosomal recessive familial Parkinson's disease. Here, we show that Parkin binds to the E2 ubiquitin-conjugating human enzyme 8 (UbcH8) through its C-terminal ring-finger. Parkin has ubiquitin-protein ligase activity in the presence of UbcH8. Parkin also ubiquitinates itself and promotes its own degradation. We also identify and show that the synaptic vesicle-associated protein, CDCrel-1, interacts with Parkin through its ring-finger domains. Furthermore, Parkin ubiquitinates and promotes the degradation of CDCrel-1. Familial-linked mutations disrupt the ubiquitin-protein ligase function of Parkin and impair Parkin and CDCrel-1 degradation. These results suggest that Parkin functions as an E3 ubiquitin-protein ligase through its ring domains and that it may control protein levels via ubiquitination. The loss of Parkin's ubiquitin-protein ligase function in familial-linked mutations suggests that this may be the cause of familial autosomal recessive Parkinson's disease.
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data-image-width="422"
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data-pmc-id="PMC27228"
data-figure-id="F1">
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id="article-image-0"
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<strong class="figure-label">
<p> Figure 1 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin interacts with the E2… </p></div>
</div>
</div>
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
<strong class="figure-label">
<p> Figure 1 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin interacts with the E2 ubiquitin-conjugating enzyme, UbcH8. ( <i> A </i> ) Multiple sequence… </p></div>
</div>
<figcaption id="figure-caption-0" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
<strong class="figure-label">
Figure 1
</strong>
<div class="figure-caption-contents">Parkin interacts with the E2 ubiquitin-conjugating enzyme, UbcH8.
(<i>A</i>) Multiple sequence alignment of ring-finger domains
of Parkin (BAA25751), HHARI (NP005735), H7-AP1 (CAB45870), and UIP28
(AAD24572). Conserved residues are highlighted, and the ring fingers
(R1 and R2) and the IBR domain are indicated. Abbreviations: HHARI,
ariadne homologue (<i>Homo sapiens</i>); H7-AP1, UbcH7-binding
protein; UIP28, UbcM4 interacting protein 28. (<i>B</i>)
Lysates prepared from HEK 293 cells transfected with pRK5-myc-Parkin
and pRK5-HAUbcH5, pRK5-HAUbcH7, or pRK5-HAUbcH8 were subjected to
immunoprecipitation (IP) with anti-myc antibody followed by anti-HA
immunoblotting. This experiment was replicated three times with similar
results.</div>
</figcaption>
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data-image-width="430"
data-image-height="407"
data-image-alt="Figure 2"
data-pmc-id="PMC27228"
data-figure-id="F2">
<img class="figure-thumb" itemprop="thumbnail"
id="article-image-1"
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alt="Figure 2" />
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<div class="figure-caption figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="caption" aria-hidden="true">
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<strong class="figure-label">
<p> Figure 2 </p>
</strong>
<div class="figure-caption-contents"><p> UbcH8 interacts preferentially with the… </p></div>
</div>
</div>
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
<strong class="figure-label">
<p> Figure 2 </p>
</strong>
<div class="figure-caption-contents"><p> UbcH8 interacts preferentially with the C-terminal R2 ring-finger domain of Parkin. ( <i> A… </i> </p></div>
</div>
<figcaption id="figure-caption-1" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
<strong class="figure-label">
Figure 2
</strong>
<div class="figure-caption-contents">UbcH8 interacts preferentially with the C-terminal R2 ring-finger
domain of Parkin. (<i>A</i>) Schematic representation of
putative functional domains of Parkin used in the mapping experiments.
(<i>B</i>) Lysates prepared from HEK 293 cells transfected with
pRK5-HA-UbcH8 and various Myc-tagged Parkin domain constructs were
subjected to immunoprecipitation (IP) with anti-myc antibody followed
by anti-HA immunoblotting (<i>Upper</i>). The blot was stripped
and reprobed with the anti-myc antibody (<i>Lower</i>) to
illustrate that equivalent levels of the Parkin constructs were present
in the extracts. This experiment was replicated three times with
similar results. (<i>C</i>) Lysates prepared from HEK 293 cells
transfected with pRK5-HA-UbcH8 and various Myc-tagged Parkin mutant
constructs (see text for details) were subjected to immunoprecipitation
(IP) with anti-myc antibody followed by anti-HA immunoblotting
(<i>Upper</i>). The blot was stripped and reprobed with the
anti-Myc antibody (<i>Lower</i>) to illustrate that equivalent
levels of the Parkin constructs were present in the extracts. This
experiment was replicated three times with similar results.</div>
</figcaption>
</figure>
<figure class="figure-item "
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data-image-width="417"
data-image-height="503"
data-image-alt="Figure 3"
data-pmc-id="PMC27228"
data-figure-id="F3">
<img class="figure-thumb" itemprop="thumbnail"
id="article-image-2"
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<div class="figure-caption figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="caption" aria-hidden="true">
<div class="caption-wrap">
<strong class="figure-label">
<p> Figure 3 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin is involved in E2-dependent… </p></div>
</div>
</div>
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
<strong class="figure-label">
<p> Figure 3 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin is involved in E2-dependent self-ubiquitination both <i> in vitro </i> and <i> in vivo </i> .… </p></div>
</div>
<figcaption id="figure-caption-2" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
<strong class="figure-label">
Figure 3
</strong>
<div class="figure-caption-contents">Parkin is involved in E2-dependent self-ubiquitination both <i>in
vitro</i> and <i>in vivo</i>. (<i>A</i>) <i>In
vitro</i> translated Parkin (5 μl) was incubated with 100 ng of
E1, 10 μg of ubiquitin (Ub) with 5 μl of bacterially expressed
UbcH8, UbcH7, UbH5, or control bacteria lysate in the ubiquitination
reaction buffer for 1.5 h at 30°C. The reaction products were
immunoblotted with anti-ubiquitin antibody. Molecular weight markers
are indicated. This experiment was replicated three times with similar
results. (<i>B</i>) <i>In vitro</i> translated Parkin
(5 μl) or Parkin mutants were incubated with 100 ng of E1, 10 μg of
Ub with 5 μl of bacterially expressed UbcH8 in the ubiquitination
reaction buffer for 1.5 h at 30°C. The reaction products were
immunoblotted with anti-ubiquitin antibody. Molecular weight markers
are indicated. This experiment was replicated three times with similar
results. (<i>C</i>) Lysates prepared from HEK 293 cells
transfected with pRK5-myc-Parkin or myc-tagged mutant Parkin constructs
and pMT123-HA-ubiquitin were subjected to immunoprecipitation (IP) with
anti-myc antibody followed by anti-HA immunoblotting
(<i>Upper</i>). The blot was stripped and reprobed with the
anti-myc antibody (<i>Lower</i>) to illustrate that equivalent
levels of the Parkin constructs were present in the extracts. This
experiment was replicated three times with similar results.
(<i>D</i>) Familial-linked Parkin mutants impair Parkin
degradation. HEK 293 cells were transfected with pRK5-myc-Parkin,
pRK5-myc-ParkinThr240Arg, pRK5-myc-Parkin-Thr415Asn, and
pRK5-myc-ParkinThr240Arg/Thr415Asn plasmids using Lipofectamine.
24 h after transfection, cells were washed and incubated with
methionine-free medium for 1 h. The cells were pulsed-chased with
100 μCi of [<sup>35</sup>S]methionine and were harvested at the
indicated times for immunoprecipitation with anti-myc antibody. The
immunoprecipitates were resolved on a 10% SDS/PAGE gel and
visualized and quantitated with a phosphoimager. This experiment was
replicated with similar results.</div>
</figcaption>
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itemprop="contentUrl"
aria-describedby="figure-caption-3"
role="button"
data-image-width="424"
data-image-height="579"
data-image-alt="Figure 4"
data-pmc-id="PMC27228"
data-figure-id="F4">
<img class="figure-thumb" itemprop="thumbnail"
id="article-image-3"
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<strong class="figure-label">
<p> Figure 4 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin interacts with, ubiquitinates and… </p></div>
</div>
</div>
<div class="figure-caption-medium figure-caption-text" aria-hidden="true">
<strong class="figure-label">
<p> Figure 4 </p>
</strong>
<div class="figure-caption-contents"><p> Parkin interacts with, ubiquitinates and promotes the degradation of CDCrel-1. ( <i> A </i> )… </p></div>
</div>
<figcaption id="figure-caption-3" class="figure-caption-full figure-caption-text" itemtype="http://schema.org/ImageObject" itemprop="description">
<strong class="figure-label">
Figure 4
</strong>
<div class="figure-caption-contents">Parkin interacts with, ubiquitinates and promotes the degradation of
CDCrel-1. (<i>A</i>) Lysates prepared from HEK 293 cells
transfected with pRK5-HA-CDCrel-1 and pRK5-myc-Parkin constructs were
subjected to immunoprecipitation (IP) with anti-myc antibody followed
by anti-HA immunoblotting. This experiment was replicated three times
with similar results. (<i>B</i>) The ring-finger domains of
Parkin interact with CDCrel-1. Lysates prepared from HEK 293 cells
transfected with pRK5-HA-CDCrel-1 and myc-tagged Parkin domain
constructs were subjected to immunoprecipitation (IP) with anti-myc
antibody followed by anti-HA immunoblotting. This experiment was
replicated three times with similar results. (<i>C</i>) Parkin
ubiquitinates CDCrel-1. HEK 293 cells were transfected with
pRK5-Myc-CDCrel-1, pMT123-HA-ubiquitin and pCMV-Parkin or control
plasmid. Cells were then treated with 1 or 2 μM clasto-Lactacystin
β-lactone for indicated times. Lysates were prepared from these cells
and subjected to immunoprecipitation (IP) with anti-myc antibody
followed by anti-HA immunoblotting. This experiment was replicated with
similar results. (<i>D</i>) Parkin promotes the degradation of
CDCrel-1. HEK 293 cells were transfected pRK5-myc-CDCrel-1 and
pRK5-myc-Parkin or empty vector. 24 h after transfection, cells
were washed and incubated with methionine-free medium for 1 h. The
cells were pulsed-chased with 100 μCi of
[<sup>35</sup>S]methionine and were harvested at the indicated times
for immunoprecipitation with anti-myc antibody. The immunoprecipitates
were resolved on a 10% SDS/PAGE gel and visualized and quantitated
with a phosphoimager. This experiment was replicated with similar
results. (<i>E</i>) Familial-linked Parkin mutants fail to
degrade CDCrel-1. The same experimental paradigm as described in Fig.
4<i>D</i> were performed with the familial-linked Parkin mutant
pRK5-myc-Parkin-Thr415Asn or pRK5-myc-ParkinGln311 and compared with
wild-type Parkin. This experiment was replicated with similar
results.</div>
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