Wu Enhui, Qiao Liang*
Ma'aikatar Chemistry, Jami'ar Fudan, Shanghai 200433, Sin
Kwayoyin cuta suna da alaƙa da cututtukan ɗan adam da lafiya. Yadda za a fahimci abubuwan da ke tattare da ƙananan ƙwayoyin cuta da ayyukansu babban batu ne da ke buƙatar yin nazari cikin gaggawa. A cikin 'yan shekarun nan, metaproteomics ya zama muhimmiyar hanyar fasaha don nazarin abun da ke ciki da aikin ƙwayoyin cuta. Koyaya, saboda sarƙaƙƙiya da babban nau'ikan samfuran al'umma na ƙayyadaddun ƙayyadaddun ƙayyadaddun ƙayyadaddun ƙayyadaddun ƙayyadaddun ƙayyadaddun samfuran al'umma, sarrafa samfuri, karɓar bayanan spectrometry da ƙididdigar bayanai sun zama manyan ƙalubale guda uku waɗanda metaproteomics ke fuskanta a halin yanzu. A cikin nazarin metaproteomics, sau da yawa ya zama dole don inganta pretreatment na nau'ikan samfurori daban-daban da kuma ɗaukar nau'ikan rarrabuwa na microbial daban-daban, haɓakawa, hakar da tsarin lysis. Kama da proteome na nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i) yana samuwa a cikin metaproteomics. Yanayin sayen bayanan DIA na iya tattara bayanan peptide na samfurin gaba ɗaya kuma yana da babban ƙarfin ci gaba. Koyaya, saboda sarkar samfuran metaproteome, binciken bayanan DIA ɗin sa ya zama babbar matsala wacce ke hana zurfin ɗaukar matakan metaproteomics. Dangane da nazarin bayanai, mataki mafi mahimmanci shi ne gina tsarin bayanan furotin. Girman girma da cikar bayanan ba wai kawai suna da tasiri mai yawa akan adadin abubuwan ganowa ba, har ma suna shafar bincike a nau'ikan nau'ikan da matakan aiki. A halin yanzu, ma'auni na zinariya don gina bayanan metaproteome shine bayanan jerin sunadaran gina jiki bisa metagenome. A lokaci guda, hanyar tace bayanan jama'a dangane da bincike na maimaitawa an kuma tabbatar da cewa yana da fa'ida mai ƙarfi mai ƙarfi. Daga hangen ƙayyadaddun dabarun nazarin bayanai, hanyoyin nazarin bayanan DIA mai tushen peptide sun mamaye cikakkiyar al'ada. Tare da haɓaka zurfin koyo da hankali na wucin gadi, zai haɓaka daidaito, ɗaukar hoto da saurin bincike na ƙididdigar macroproteomic. Dangane da bincike na bioinformatics na ƙasa, an haɓaka jerin kayan aikin annotation a cikin 'yan shekarun nan, waɗanda za su iya yin bayanin nau'ikan nau'ikan nau'ikan furotin, matakin peptide da matakin ƙwayar cuta don samun abubuwan da ke tattare da ƙananan ƙwayoyin cuta. Idan aka kwatanta da sauran hanyoyin omics, nazarin ayyuka na al'ummomin ƙananan ƙwayoyin cuta wani siffa ce ta musamman na macroproteomics. Macroproteomics ya zama wani muhimmin ɓangare na bincike na multi-omics na al'ummomin ƙananan ƙwayoyin cuta, kuma har yanzu yana da babban ƙarfin ci gaba dangane da zurfin ɗaukar hoto, ganewar ganewa, da cikakkun bayanai.
01 Misalin pretreatment
A halin yanzu, ana amfani da fasahar metaproteomics sosai a cikin binciken microbiome na ɗan adam, ƙasa, abinci, teku, sludge mai aiki da sauran filayen. Idan aka kwatanta da binciken proteome na nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'in nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i nau'i)), samfurin pretreatment na metaproteome na samfurori masu rikitarwa yana fuskantar ƙarin kalubale. Abubuwan da ke cikin ƙananan ƙwayoyin cuta a cikin samfurori na ainihi suna da wuyar gaske, kewayon haɓaka mai yawa yana da girma, tsarin bangon tantanin halitta na nau'ikan ƙwayoyin cuta daban-daban ya bambanta sosai, kuma samfuran sau da yawa sun ƙunshi babban adadin furotin mai masauki da sauran ƙazanta. Sabili da haka, a cikin nazarin metaproteome, sau da yawa ya zama dole don inganta nau'ikan samfurori daban-daban da kuma ɗaukar nau'ikan rarrabuwa daban-daban, haɓakawa, haɓakawa da tsarin lysis.
Fitar da ƙananan ƙwayoyin metaproteomes daga samfurori daban-daban yana da wasu kamanceceniya da kuma wasu bambance-bambance, amma a halin yanzu akwai rashin ingantaccen tsarin aiwatarwa don nau'ikan samfuran metaproteome daban-daban.
02Mass spectrometry data samu
A cikin bincike na proteome na harbi, cakuda peptide bayan pretreatment an fara rabu da shi a cikin ginshiƙi na chromatographic, sannan ya shiga cikin ma'auni na taro don samun bayanai bayan ionization. Hakanan jinsi guda na asali na asali, taro na bayanan da aka samu a cikin bincike na Macroprootote ya haɗa da DDA yanayin da yanayin Diaallin.
Tare da ci gaba da haɓakawa da sabuntawa na kayan aikin gani na taro, ana amfani da na'urori masu yawan gaske tare da mafi girman hankali da ƙuduri zuwa metaproteome, kuma zurfin ɗaukar hoto na binciken metaproteome shima yana ci gaba da inganta. Na dogon lokaci, an yi amfani da jerin manyan kayan aikin na'ura mai ƙarfi wanda Orbitrap ke jagoranta a cikin metaproteome.
Teburin 1 na ainihin rubutun yana nuna wasu nazarin wakilai akan metaproteomics daga 2011 zuwa yanzu dangane da nau'in samfurin, dabarun bincike, kayan aikin ƙira, hanyar saye, software na bincike, da adadin ganowa.
03 Mass Spectrometry Data Anan
3.1 Dabarun nazarin bayanan DDA
3.1.1 Binciken Database
3.1.2da novodabarun jeri
3.2 Dabarun nazarin bayanan DIA
04 Rarraba nau'ikan nau'ikan da bayanin aiki
Ƙirƙirar al'ummomin ƙananan ƙwayoyin cuta a matakan haraji daban-daban shine ɗayan mahimman wuraren bincike a cikin binciken microbiome. A cikin 'yan shekarun nan, an ƙirƙiri jerin kayan aikin ƙididdiga don bayyana nau'in nau'in nau'in nau'in furotin, matakin peptide, da matakin kwayoyin halitta don samun abubuwan da ke tattare da ƙananan ƙwayoyin cuta.
Ma'anar bayanin aiki shine kwatanta jerin abubuwan furotin da aka yi niyya tare da bayanan jerin sunadaran aiki. Yin amfani da bayanan ayyukan kwayoyin halitta kamar GO, COG, KEGG, eggNOG, da dai sauransu, ana iya yin nazarin bayanan aikin daban-daban akan sunadaran da macroproteomes suka gano. Kayan aikin bayanin sun haɗa da Blast2GO, DAVID, KOBAS, da sauransu.
05Summary da Outlook
Ƙananan ƙwayoyin cuta suna taka muhimmiyar rawa a lafiyar ɗan adam da cututtuka. A cikin 'yan shekarun nan, metaproteomics ya zama muhimmiyar hanyar fasaha don nazarin aikin al'ummomin ƙananan ƙwayoyin cuta. Tsarin nazari na metaproteomics yayi kama da na nau'ikan nau'ikan proteomics guda ɗaya, amma saboda rikitarwa na abin bincike na metaproteomics, takamaiman dabarun bincike suna buƙatar ɗaukar matakan bincike a kowane matakin bincike, daga samfurin pretreatment, samun bayanai zuwa nazarin bayanai. A halin yanzu, godiya ga inganta hanyoyin da aka rigaya, da ci gaba da haɓaka fasahar fasaha mai yawa da kuma saurin ci gaban bioinformatics, metaproteomics ya sami babban ci gaba a cikin zurfin ganewa da iyakokin aikace-aikace.
A cikin aiwatar da pre-jiyya na macroproteome samfurori, dole ne a fara la'akari da yanayin samfurin. Yadda za a raba ƙananan ƙwayoyin cuta daga ƙwayoyin muhalli da sunadaran suna ɗaya daga cikin manyan ƙalubalen da ke fuskantar macroproteomes, kuma daidaito tsakanin ingancin rabuwa da asarar ƙananan ƙwayoyin cuta matsala ce ta gaggawa da za a warware. Na biyu, hakar sunadaran ƙwayoyin cuta dole ne a yi la'akari da bambance-bambancen da ke haifar da yanayin tsarin ƙwayoyin cuta daban-daban. Samfuran macroproteome a cikin kewayon alama kuma suna buƙatar takamaiman hanyoyin magani kafin magani.
Dangane da na'urorin na'ura mai ba da hanya tsakanin hanyoyin sadarwa, na'urori na al'ada na al'ada sun sami canji daga na'urori masu yawa dangane da masu nazarin taro na Orbitrap kamar LTQ-Orbitrap da Q Exactive to mass spectrometers bisa la'akari da motsin ion haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe-haɗe. . Jerin timsTOF na kayan aiki tare da bayanin girman motsin ion suna da ingantaccen ganowa, ƙarancin ganowa, da ingantaccen maimaitawa. A hankali sun zama kayan aiki masu mahimmanci a fagagen bincike iri-iri waɗanda ke buƙatar gano yawan abubuwan gani, kamar su proteome, metaproteome, da metabolome na nau'in nau'i ɗaya. Yana da kyau a lura cewa na dogon lokaci, ƙwaƙƙwaran kewayon na'urorin spectrometry na taro ya iyakance zurfin ɗaukar hoto na binciken metaproteome. A nan gaba, na'urori masu yawan gaske tare da babban kewayon kuzari na iya haɓaka hankali da daidaiton gano furotin a cikin metaproteomes.
Don sayan bayanan yawan jama'a, kodayake yanayin sayan bayanan DIA ya kasance cikin karbuwa sosai a cikin tsarin sinadarai guda ɗaya, yawancin nazarin macroproteome na yanzu har yanzu suna amfani da yanayin sayan bayanan DDA. Yanayin sayan bayanan DIA zai iya samun cikakkiyar bayanan guntun ion na samfurin, kuma idan aka kwatanta da yanayin sayan bayanan DDA, yana da yuwuwar samun cikakken bayanin peptide na samfurin macroproteome. Duk da haka, saboda babban rikitarwa na bayanan DIA, nazarin bayanan macroproteome na DIA har yanzu yana fuskantar manyan matsaloli. Ana sa ran haɓaka ƙwarewar wucin gadi da zurfin koyo don inganta daidaito da cikar nazarin bayanan DIA.
A cikin nazarin bayanan metaproteomics, ɗaya daga cikin mahimman matakai shine gina bayanan bayanan furotin. Don shahararrun wuraren bincike irin su flora na hanji, ana iya amfani da bayanan ƙananan ƙwayoyin cuta na hanji kamar IGC da HMP, kuma an sami sakamako mai kyau na ganewa. Ga mafi yawan sauran nazarin metaproteomics, dabarun gina bayanai mafi inganci har yanzu shine a kafa takamammen takamaiman bayanan sunadaran sunadarai dangane da bayanan jeri na metagenomic. Don samfuran al'ummomin microbialai tare da babban rikitarwa da kuma kewayon ƙarfi, wajibi ne don ƙara zurfin samfuran ƙananan nau'ikan bayanan. Lokacin da aka rasa jerin bayanai, ana iya amfani da hanyar bincike mai maimaitawa don inganta bayanan jama'a. Duk da haka, bincike na maimaitawa na iya shafar sarrafa ingancin FDR, don haka ana buƙatar bincika sakamakon binciken a hankali. Bugu da kari, amfani da tsarin kula da ingancin ingancin FDR na gargajiya a cikin binciken metaproteomics har yanzu yana da daraja a bincika. Dangane da dabarun bincike, dabarun ɗakin karatu na ƙawance na iya haɓaka zurfin ɗaukar hoto na metaproteomics DIA. A cikin 'yan shekarun nan, ɗakin karatu da aka annabta da aka ƙirƙira bisa zurfafa ilmantarwa ya nuna kyakkyawan aiki a cikin abubuwan haɓakar DIA. Koyaya, ma'ajin bayanai na metaproteome galibi suna ɗauke da miliyoyin shigarwar sunadaran gina jiki, wanda ke haifar da babban sikelin dakunan karatu da aka annabta, suna cinye albarkatu masu yawa, kuma suna haifar da babban filin bincike. Bugu da kari, kamanceceniya tsakanin jerin furotin a cikin metaproteomes ya bambanta sosai, yana sa yana da wahala a tabbatar da daidaiton ƙirar tsinkayar ɗakin karatu, don haka ba a yi amfani da ɗakunan karatu da aka annabta ba a cikin metaproteomics. Bugu da kari, ana buƙatar samar da sabbin dabarun gina jiki da dabarun tantance rabe-rabe don amfani da nazarin metaproteomics na sunadaran sunadaran jeri sosai.
A taƙaice, a matsayin fasahar bincike na microbiome mai tasowa, fasahar metaproteomics ta sami sakamako mai mahimmanci na bincike kuma tana da babban ƙarfin ci gaba.
Lokacin aikawa: Agusta-30-2024