Wu Enhui, Qiao Liang*
Keʻena ʻOihana Kimia, Ke Kulanui ʻo Fudan, Shanghai 200433, Kina
Pili nā microorganisms i nā maʻi a me ke olakino. Pehea e hoʻomaopopo ai i ka hoʻokumu ʻana o nā kaiāulu microbial a me kā lākou mau hana he pilikia nui e pono e aʻo koke ʻia. I nā makahiki i hala iho nei, ua lilo ka metaproteomic i mea ʻenehana koʻikoʻi e aʻo ai i ka haku mele a me ka hana o nā microorganism. Eia nō naʻe, ma muli o ka paʻakikī a me ka heterogeneity kiʻekiʻe o nā microbial community samples, sample processing, mass spectrometry data acquisition and data analysis i lilo i ʻekolu mau pilikia nui i kēia manawa e ka metaproteomics. I loko o ka metaproteomics analysis, pono pinepine e hoʻomaʻamaʻa i ka pretreatment o nā ʻano laʻana like ʻole a hoʻohana i nā ʻano like ʻole o ka microbial separation, enrichment, extraction and lysis schemes. E like me ka proteome o kahi ʻano hoʻokahi, ʻo nā ʻano ʻano ʻikepili kikoʻī kikoʻī nui i loko o nā metaproteomics e pili ana i ka mode data-dependent acquisition (DDA) a me ka data-independent acquisition (DIA). Hiki i ke ʻano o ka loaʻa ʻana o ka ʻikepili DIA ke hōʻiliʻili piha i ka ʻike peptide o ka laʻana a loaʻa iā ia ka mana hoʻomohala nui. Eia naʻe, ma muli o ka paʻakikī o nā laʻana metaproteome, ua lilo kāna ʻikepili DIA i pilikia nui e keʻakeʻa ana i ka uhi hohonu o nā metaproteomics. Ma ke ʻano o ka nānā ʻana i ka ʻikepili, ʻo ka hana koʻikoʻi ke kūkulu ʻana i kahi waihona sequence protein. ʻO ka nui a me ka hoʻopiha piha ʻana o ka waihona ʻaʻole wale ka hopena nui i ka helu o nā ʻike, akā pili pū kekahi i ka nānā ʻana i nā ʻano a me nā pae hana. I kēia manawa, ʻo ke kūlana gula no ke kūkulu ʻana i kahi waihona metaproteome he waihona kaʻina hana protein e pili ana i ka metagenome. Ma ka manawa like, ua hōʻike ʻia ke ʻano kānana ʻikepili lehulehu e pili ana i ka huli hoʻololi ʻana i ka waiwai kūpono. Mai ka hiʻohiʻona o nā hoʻolālā ʻikepili kikoʻī kikoʻī, ua noho nā ʻano loiloi data DIA i ka peptide-centered i kahi kumu nui. Me ka hoʻomohala ʻana o ke aʻo hohonu a me ka naʻauao hana, e hoʻolaha nui ia i ka pololei, ka uhi a me ka wikiwiki o ka nānā ʻana o ka ʻikepili macroproteomic data. Ma ke ʻano o ka loiloi bioinformatics i lalo, ua hoʻomohala ʻia kahi ʻano o nā mea hana annotation i nā makahiki i hala iho nei, hiki ke hana i ka annotation species ma ka pae protein, pae peptide a me ka pae gene e loaʻa ai ka haku o nā kaiāulu microbial. Ke hoʻohālikelike ʻia me nā ʻano omics ʻē aʻe, ʻo ka loiloi hana o nā kaiāulu microbial kahi hiʻohiʻona kūʻokoʻa o macroproteomics. Ua lilo ʻo Macroproteomics i ʻāpana koʻikoʻi o ka loiloi multi-omics o nā kaiāulu microbial, a loaʻa mau ka mana hoʻomohala nui ma ke ʻano o ka hohonu o ka uhi, ka ʻike ʻike, a me ka piha ʻana o ka ʻikepili.
01Laʻana hana mua
I kēia manawa, ua hoʻohana nui ʻia ka ʻenehana metaproteomics i ka noiʻi ʻana i ka microbiome kanaka, ka lepo, ka meaʻai, ka moana, ka sludge ikaika a me nā māla ʻē aʻe. Ke hoʻohālikelike ʻia me ka nānā ʻana o ka proteome o kahi ʻano hoʻokahi, ʻoi aku ka nui o nā pilikia o ka laʻana o ka metaproteome o nā laʻana paʻakikī. He paʻakikī ke ʻano microbial i loko o nā laʻana maoli, nui ka nui o ka nui, ʻokoʻa ke ʻano o ka pā hale o nā ʻano microorganisms like ʻole, a loaʻa pinepine nā mea hoʻohālike i ka nui o nā protein host a me nā mea haumia ʻē aʻe. No laila, i ka nānā ʻana i ka metaproteome, pono pinepine e hoʻonui i nā ʻano laʻana like ʻole a hoʻohana i nā ʻano like ʻole o ka microbial separation, enrichment, extraction a me lysis schemes.
ʻO ka unuhi ʻana o nā microbial metaproteomes mai nā laʻana like ʻole he mau mea like a me kekahi mau ʻokoʻa, akā i kēia manawa aia ka nele o kahi kaʻina hana mua i hui ʻia no nā ʻano like ʻole o nā ʻano metaproteome.
02Ka loaʻa ʻana o ka ʻikepili spectrometry
I ka panapana panapana proteome, ua hoʻokaʻawale mua ʻia ka hui peptide ma hope o ka hoʻomaʻamaʻa mua ʻana i ke kolamu chromatographic, a laila komo i ka spectrometer nui no ka loaʻa ʻana o ka ʻikepili ma hope o ka ionization. E like me ka nānā ʻana i ka proteome ʻano ʻano hoʻokahi, ʻo nā ʻano ʻano loaʻa ʻikepili spectrometry nui i ka nānā ʻana macroproteome e loaʻa i ke ʻano DDA a me ke ʻano DIA.
Me ka hoʻomau mau ʻana a me ka hōʻano hou ʻana o nā mea hana spectrometry nuipa, hoʻohana ʻia nā mea hana spectrometry me ka ʻike kiʻekiʻe a me ka hoʻonā i ka metaproteome, a ua hoʻomaikaʻi mau ʻia ka hohonu o ka uhi ʻana o ka metaproteome analysis. No ka manawa lōʻihi, ua hoʻohana nui ʻia kahi pūʻulu o nā mea hana kiʻekiʻe kiʻekiʻe e alakaʻi ʻia e Orbitrap i metaproteome.
Hōʻike ka papa 1 o ka kikokikona kumu i kekahi mau haʻawina e pili ana i nā metaproteomics mai 2011 a hiki i kēia manawa e pili ana i ke ʻano o ka laʻana, ka hoʻolālā ʻikepili, ka mea hana spectrometry lehulehu, ke ʻano o ka loaʻa ʻana, ka polokalamu loiloi, a me ka helu o nā ʻike.
03Ka ʻikepili ʻike kikoʻī nui
3.1 Ka hoʻolālā ʻikepili DDA
3.1.1 Huli waihona
3.1.2de novohoʻolālā hoʻonohonoho
3.2 Hoʻolālā ʻikepili ʻikepili DIA
04 Ka hoʻokaʻawale ʻana o nā ʻano a me ka wehewehe hana
ʻO ka hui ʻana o nā kaiāulu microbial ma nā pae ʻauhau ʻokoʻa kekahi o nā wahi noiʻi nui i ka noiʻi microbiome. I nā makahiki i hala iho nei, ua hoʻomohala ʻia kahi pūʻulu o nā mea hana annotation e hōʻike i nā ʻano ma ka pae protein, pae peptide, a me ka pae gene e loaʻa ai ke ʻano o nā kaiāulu microbial.
ʻO ke kumu o ka palapala hoʻopuka hana ʻo ia ka hoʻohālikelike ʻana i ke kaʻina o ka protein target me ka database sequence protein functional. Ke hoʻohana nei i nā ʻikepili hana gene e like me GO, COG, KEGG, eggNOG, a me nā mea ʻē aʻe, hiki ke hoʻokō ʻia nā loiloi annotation hana like ʻole ma nā protein i ʻike ʻia e nā macroproteomes. ʻO nā mea hana hōʻike me Blast2GO, DAVID, KOBAS, etc.
05 Hōʻuluʻulu manaʻo a me ka nānā
He kuleana koʻikoʻi nā microorganism i ke olakino a me nā maʻi kanaka. I nā makahiki i hala iho nei, ua lilo ka metaproteomics i mea ʻenehana koʻikoʻi e aʻo ai i ka hana o nā kaiāulu microbial. Ka analytical kaʻina hana o metaproteomics ua like ia me ka hookahi-species proteomics, akā, ma muli o ka paʻakikī o ka noiʻi mea o metaproteomics, kiko'ī noiʻi hoʻolālā e pono e aponoia i loko o kēlā me kēia kaʻina kālailai, mai laʻana pretreatment,ʻikepili acquisition i ka ikepili kālailai. I kēia manawa, mahalo i ka hoʻomaikaʻi ʻana i nā ʻano hana pretreatment, ka hoʻomau mau ʻana o ka ʻenehana spectrometry nui a me ka hoʻomohala wikiwiki ʻana o nā bioinformatics, ua holomua nui ka metaproteomics i ka hohonu o ka ʻike a me ka pae noi.
I ke kaʻina o ka hoʻomaʻamaʻa mua ʻana i nā laʻana macroproteome, pono e noʻonoʻo mua ke ʻano o ka hāpana. ʻO ke ʻano o ka hoʻokaʻawale ʻana i nā microorganism mai nā cell kaiapuni a me nā protein kekahi o nā pilikia koʻikoʻi e kū nei i nā macroproteomes, a ʻo ke kaulike ma waena o ka hoʻokaʻawale ʻana a me ka nalowale microbial he pilikia koʻikoʻi e hoʻoponopono ʻia. ʻO ka lua, pono e noʻonoʻo ka lawe ʻana i ka protein o nā microorganism i nā ʻokoʻa i hoʻokumu ʻia e ka heterogeneity structural o nā bacteria like ʻole. Pono pū nā laʻana macroproteome i ka laʻana i nā ʻano hana mua o ka mālama ʻana.
Ma ke ʻano o nā mea hana spectrometry nui, ua hoʻololi ʻia nā mea hana spectrometry nui mai nā spectrometers nui i hoʻokumu ʻia ma Orbitrap mass analyrs e like me LTQ-Orbitrap a me Q Exactive i nā spectrometers nui ma muli o ka mobility ion i hui pū ʻia nā mea ananaly time-of-flight e like me timsTOF Pro. . ʻO ka pūʻulu timsTOF o nā mea kani me ka ʻike ion mobility dimension he kiʻekiʻe ka ʻike pololei, haʻahaʻa ka palena ʻike, a me ka hiki ke hana hou. Ua lilo lākou i mau mea hana koʻikoʻi i nā ʻano noiʻi like ʻole e koi ana i ka ʻike kikoʻī nui, e like me ka proteome, metaproteome, a me ka metabolome o kahi ʻano hoʻokahi. Pono e hoʻomaopopo ʻia no ka manawa lōʻihi, ua kaupalena ka laulā o nā mea hana spectrometry i ka hohonu o ka uhi protein o ka noiʻi metaproteome. I ka wā e hiki mai ana, hiki i nā mea hana spectrometry nui me ka laulā ikaika nui ke hoʻomaikaʻi i ka ʻike a me ka pololei o ka ʻike protein i nā metaproteomes.
No ka loaʻa ʻana o ka ʻikepili spectrometry nui, ʻoiai ua hoʻohana nui ʻia ke ʻano o ka loaʻa ʻana o ka ʻikepili DIA i loko o ka proteome o kahi ʻano hoʻokahi, ʻo ka hapa nui o nā loiloi macroproteome o kēia manawa ke hoʻohana nei i ke ʻano ʻikepili DDA. Hiki ke loa'a piha i ke 'ano o ka loa'a 'ikepili DIA i ka 'ikepili ion o ka hāpana, a i ka ho'ohālikelike 'ia me ke 'ano o ka loa'a 'ikepili DDA, hiki ke loa'a ka 'ike peptide o ka hāpana macroproteome. Eia nō naʻe, ma muli o ka paʻakikī kiʻekiʻe o ka ʻikepili DIA, ke kū nei ka loiloi o ka ʻikepili macroproteome DIA i nā pilikia nui. Manaʻo ʻia ka hoʻomohala ʻana o ka naʻauao akamai a me ka hoʻonaʻauao hohonu e hoʻomaikaʻi i ka pololei a me ka hoʻopiha piha ʻana o ka ʻikepili ʻikepili DIA.
I ka ʻikepili ʻikepili o metaproteomics, ʻo kekahi o nā ʻanuʻu koʻikoʻi ke kūkulu ʻana i ka ʻikepili sequence protein. No nā wahi noiʻi kaulana e like me ka flora intestinal, hiki ke hoʻohana ʻia nā ʻikepili microbial intestinal e like me IGC a me HMP, a ua loaʻa nā hopena ʻike maikaʻi. No ka hapanui o nā kālailai metaproteomics ʻē aʻe, ʻo ka hoʻolālā kūkulu ʻikepili ʻoi aku ka maikaʻi o ka hoʻokumu ʻana i kahi hōkeo pūmua protein kikoʻī kikoʻī e pili ana i ka ʻikepili sequencing metagenomic. No nā laʻana kaiaulu microbial me ka paʻakikī kiʻekiʻe a me ka laulā dynamic nui, pono e hoʻonui i ka hohonu o ke kaʻina hana e hoʻonui i ka ʻike ʻana i nā ʻano haʻahaʻa haʻahaʻa, a laila e hoʻomaikaʻi ai i ka uhi ʻana o ka waihona sequence protein. Ke nele ka ʻikepili helu, hiki ke hoʻohana ʻia kahi ʻano hulina e hoʻohana i ka ʻikepili lehulehu. Eia nō naʻe, hiki i ka huli ʻana ke hoʻololi i ka mana maikaʻi FDR, no laila pono e nānā pono ʻia nā hopena hulina. Eia kekahi, ʻo ka hoʻohana ʻana o nā hiʻohiʻona mana o ka maikaʻi FDR kuʻuna i ka metaproteomics analysis he mea pono e ʻimi. Ma ke ʻano o ka hoʻolālā ʻimi, hiki i ka hoʻolālā hale waihona puke hybrid ke hoʻomaikaʻi i ka hohonu o ka uhi o DIA metaproteomics. I nā makahiki i hala iho nei, ua hōʻike ʻia ka waihona spectral i wānana ʻia e pili ana i ke aʻo hohonu i ka hana kiʻekiʻe ma DIA proteomics. Eia nō naʻe, loaʻa i nā ʻikepili metaproteome he mau miliona o nā komo pūmua, kahi i loaʻa ai ka nui o nā hale waihona puke spectral i wānana ʻia, hoʻopau i ka nui o nā kumuwaiwai helu, a loaʻa i kahi wahi huli nui. Eia kekahi, ʻokoʻa ka like ʻana ma waena o nā kaʻina protein i loko o nā metaproteomes, he mea paʻakikī ke hōʻoia i ka pololei o ka spectral library prediction model, no laila ʻaʻole i hoʻohana nui ʻia nā hale waihona spectral i wānana ʻia i nā metaproteomics. Eia kekahi, pono e hoʻomohala ʻia nā hoʻolālā hoʻolālā protein hou a me ka hoʻokaʻawale ʻana e pili i ka metaproteomics analysis o nā protein like-like.
I ka hōʻuluʻulu ʻana, ma ke ʻano he ʻenehana noiʻi microbiome e puka mai nei, ua loaʻa i ka ʻenehana metaproteomics nā hopena noiʻi koʻikoʻi a loaʻa pū kekahi i ka hiki ke hoʻomohala nui.
Ka manawa hoʻouna: ʻAukake-30-2024