{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T10:28:17Z","timestamp":1785407297899,"version":"3.56.0"},"posted":{"date-parts":[[2025,8,1]]},"group-title":"Microbiology","reference-count":42,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,8,1]],"date-time":"2025-08-01T00:00:00Z","timestamp":1754006400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"award":["R21AI187107"],"award-info":[{"award-number":["R21AI187107"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["S10OD024978"],"award-info":[{"award-number":["S10OD024978"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["R01AI114800"],"award-info":[{"award-number":["R01AI114800"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]},{"award":["P30CA013148"],"award-info":[{"award-number":["P30CA013148"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,8,1]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>\n                  <jats:italic>Streptococcus pneumoniae<\/jats:italic>\n                  is one of the most important human respiratory pathogens worldwide. The increase in antibiotic resistance in\n                  <jats:italic>S. pneumoniae<\/jats:italic>\n                  and other pathogens is a significant public health concern. The streptococcal 70S ribosome is a prime target for antibiotics. Ribosomal protein L27 reaches into the peptidyl transferase center with its extended N-terminus and may be involved in the translation process. We have shown that L27 in Firmicutes, including staphylococci and streptococci, has an additional 9-12 amino acid N-terminal extension compared to Gram-negative organisms like\n                  <jats:italic>Escherichia coli<\/jats:italic>\n                  . The extension is cleaved by a protease called Prp that is absent from organisms that lack the extension. In\n                  <jats:italic>S. aureus<\/jats:italic>\n                  , Prp and the N-terminal extension of L27 are essential.\n                <\/jats:p>\n                <jats:p>\n                  Here, we have characterized the cleavage of L27 by Prp in\n                  <jats:italic>S. pneumoniae<\/jats:italic>\n                  . Prp forms dimers that efficiently cleave L27\n                  <jats:italic>in vitro<\/jats:italic>\n                  . An inactive form of Prp (PrpC34S) binds to L27 without cleaving, whereas L27 with a mutation (F12A) of the cleavage site does not bind Prp. Overexpression of PrpC34S\n                  <jats:italic>in vivo<\/jats:italic>\n                  is detrimental to\n                  <jats:italic>S. pneumoniae<\/jats:italic>\n                  growth. Surprisingly, a\n                  <jats:italic>S. pneumoniae<\/jats:italic>\n                  \u0394\n                  <jats:italic>prp<\/jats:italic>\n                  strain was viable, apparently due to cleavage of L27 by another, unknown protease. Unlike in\n                  <jats:italic>S. aureus<\/jats:italic>\n                  , a mutant strain lacking the N-terminal extension of L27 was viable, but showed impaired growth. Our study sheds light on a process that could be exploited for novel antibiotics, but emphasizes important differences between streptococci and staphylococci.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>HIGHLIGHTS<\/jats:title>\n                  <jats:list list-type=\"bullet\">\n                    <jats:list-item>\n                      <jats:p>\n                        Ribosomal protein L27 in\n                        <jats:italic>S. pneumoniae<\/jats:italic>\n                        is N-terminally processed by Prp protease\n                      <\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>\n                        A\n                        <jats:italic>S. pneumoniae<\/jats:italic>\n                        \u0394\n                        <jats:italic>prp<\/jats:italic>\n                        mutant is viable, but exhibits impaired growth\n                      <\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Cleavage of L27 is required for viability, but the N-terminal extension is not essential<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>In the absence of Prp, L27 is processed by another protease.<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>\n                        There are distinct differences in the role of Prp between\n                        <jats:italic>S. aureus<\/jats:italic>\n                        and\n                        <jats:italic>S. pneumoniae<\/jats:italic>\n                        .\n                      <\/jats:p>\n                    <\/jats:list-item>\n                  <\/jats:list>\n                <\/jats:sec>","DOI":"10.1101\/2025.08.01.668195","type":"posted-content","created":{"date-parts":[[2025,8,2]],"date-time":"2025-08-02T04:00:20Z","timestamp":1754107220000},"source":"Crossref","is-referenced-by-count":0,"title":["Cleavage of\n                  <i>Streptococcus pneumoniae<\/i>\n                  ribosomal protein L27 by the Prp protease"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4458-3901","authenticated-orcid":false,"given":"Amarshi","family":"Mukherjee","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2056-6311","authenticated-orcid":false,"given":"Mohamed O.","family":"Nasef","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3277-8664","authenticated-orcid":false,"given":"Patrick M.","family":"Lindstrom","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Vipin","family":"Chembilikandy","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8542-4069","authenticated-orcid":false,"given":"Carlos J.","family":"Orihuela","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5655-4123","authenticated-orcid":false,"given":"Terje","family":"Dokland","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2025080513250905000_2025.08.01.668195v1.1","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro1871"},{"key":"2025080513250905000_2025.08.01.668195v1.2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1469-0691.2010.03183.x"},{"key":"2025080513250905000_2025.08.01.668195v1.3","unstructured":"Dion C.F. & Ashurst J.V. 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