Skip Header

You are using a version of browser that may not display all the features of this website. Please consider upgrading your browser.
Entry version 116 (12 Aug 2020)
Sequence version 1 (01 May 2000)
Previous versions | rss
Help videoAdd a publicationFeedback
Protein

Glucosidase 2 subunit alpha

Gene

gls2

Organism
Schizosaccharomyces pombe (strain 972 / ATCC 24843) (Fission yeast)
Status
Reviewed-Annotation score:

Annotation score:4 out of 5

<p>The annotation score provides a heuristic measure of the annotation content of a UniProtKB entry or proteome. This score <strong>cannot</strong> be used as a measure of the accuracy of the annotation as we cannot define the 'correct annotation' for any given protein.<p><a href='/help/annotation_score' target='_top'>More...</a></p>
-Experimental evidence at protein leveli <p>This indicates the type of evidence that supports the existence of the protein. Note that the 'protein existence' evidence does not give information on the accuracy or correctness of the sequence(s) displayed.<p><a href='/help/protein_existence' target='_top'>More...</a></p>

<p>This section provides any useful information about the protein, mostly biological knowledge.<p><a href='/help/function_section' target='_top'>More...</a></p>Functioni

Catalytic subunit of glucosidase 2, which cleaves sequentially the 2 innermost alpha-1,3-linked glucose residues from the Glc2Man9GlcNAc2 oligosaccharide precursor of immature glycoproteins.By similarity

<p>This subsection of the <a href="http://www.uniprot.org/help/function%5Fsection">Function</a> section describes the catalytic activity of an enzyme, i.e. a chemical reaction that the enzyme catalyzes.<p><a href='/help/catalytic_activity' target='_top'>More...</a></p>Catalytic activityi

<p>This subsection of the <a href="http://www.uniprot.org/help/function%5Fsection">'Function'</a> section describes the metabolic pathway(s) associated with a protein.<p><a href='/help/pathway' target='_top'>More...</a></p>Pathwayi: N-glycan metabolism

This protein is involved in the pathway N-glycan metabolism, which is part of Glycan metabolism.
View all proteins of this organism that are known to be involved in the pathway N-glycan metabolism and in Glycan metabolism.

Sites

Feature keyPosition(s)DescriptionActionsGraphical viewLength
<p>This subsection of the <a href="http://www.uniprot.org/help/function%5Fsection">Function</a> section is used for enzymes and indicates the residues directly involved in catalysis.<p><a href='/help/act_site' target='_top'>More...</a></p>Active sitei524NucleophileBy similarity1
Active sitei527By similarity1
Active sitei600Proton donorBy similarity1

<p>The <a href="http://www.geneontology.org/">Gene Ontology (GO)</a> project provides a set of hierarchical controlled vocabulary split into 3 categories:<p><a href='/help/gene_ontology' target='_top'>More...</a></p>GO - Molecular functioni

GO - Biological processi

<p>UniProtKB Keywords constitute a <a href="http://www.uniprot.org/keywords">controlled vocabulary</a> with a hierarchical structure. Keywords summarise the content of a UniProtKB entry and facilitate the search for proteins of interest.<p><a href='/help/keywords' target='_top'>More...</a></p>Keywordsi

Molecular functionGlycosidase, Hydrolase

Enzyme and pathway databases

UniPathway: a resource for the exploration and annotation of metabolic pathways

More...
UniPathwayi
UPA00957

Protein family/group databases

Carbohydrate-Active enZymes

More...
CAZyi
GH31, Glycoside Hydrolase Family 31

<p>This section provides information about the protein and gene name(s) and synonym(s) and about the organism that is the source of the protein sequence.<p><a href='/help/names_and_taxonomy_section' target='_top'>More...</a></p>Names & Taxonomyi

<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section provides an exhaustive list of all names of the protein, from commonly used to obsolete, to allow unambiguous identification of a protein.<p><a href='/help/protein_names' target='_top'>More...</a></p>Protein namesi
Recommended name:
Glucosidase 2 subunit alphaBy similarity (EC:3.2.1.2072 Publications)
Alternative name(s):
Alpha-glucosidase II subunit alphaBy similarity
Glucosidase II gls2Imported
Glucosidase II subunit alphaBy similarity
<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section indicates the name(s) of the gene(s) that code for the protein sequence(s) described in the entry. Four distinct tokens exist: 'Name', 'Synonyms', 'Ordered locus names' and 'ORF names'.<p><a href='/help/gene_name' target='_top'>More...</a></p>Gene namesi
Name:gls2Imported
ORF Names:SPAC1002.03c
<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section provides information on the name(s) of the organism that is the source of the protein sequence.<p><a href='/help/organism-name' target='_top'>More...</a></p>OrganismiSchizosaccharomyces pombe (strain 972 / ATCC 24843) (Fission yeast)
<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section shows the unique identifier assigned by the NCBI to the source organism of the protein. This is known as the 'taxonomic identifier' or 'taxid'.<p><a href='/help/taxonomic_identifier' target='_top'>More...</a></p>Taxonomic identifieri284812 [NCBI]
<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section contains the taxonomic hierarchical classification lineage of the source organism. It lists the nodes as they appear top-down in the taxonomic tree, with the more general grouping listed first.<p><a href='/help/taxonomic_lineage' target='_top'>More...</a></p>Taxonomic lineageiEukaryotaFungiDikaryaAscomycotaTaphrinomycotinaSchizosaccharomycetesSchizosaccharomycetalesSchizosaccharomycetaceaeSchizosaccharomyces
<p>This subsection of the <a href="http://www.uniprot.org/help/names%5Fand%5Ftaxonomy%5Fsection">Names and taxonomy</a> section is present for entries that are part of a <a href="http://www.uniprot.org/proteomes">proteome</a>, i.e. of a set of proteins thought to be expressed by organisms whose genomes have been completely sequenced.<p><a href='/help/proteomes_manual' target='_top'>More...</a></p>Proteomesi
  • UP000002485 <p>A UniProt <a href="http://www.uniprot.org/manual/proteomes%5Fmanual">proteome</a> can consist of several components.<br></br>The component name refers to the genomic component encoding a set of proteins.<p><a href='/help/proteome_component' target='_top'>More...</a></p> Componenti: Chromosome I

Organism-specific databases

Eukaryotic Pathogen Database Resources

More...
EuPathDBi
FungiDB:SPAC1002.03c

Schizosaccharomyces pombe database

More...
PomBasei
SPAC1002.03c, gls2

<p>This section provides information on the location and the topology of the mature protein in the cell.<p><a href='/help/subcellular_location_section' target='_top'>More...</a></p>Subcellular locationi

Extracellular region or secreted Cytosol Plasma membrane Cell wall Cytoskeleton Vacuole Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionGraphics by Christian Stolte & Seán O’Donoghue; Source: COMPARTMENTS

Keywords - Cellular componenti

Endoplasmic reticulum

<p>This section provides information on the disease(s) and phenotype(s) associated with a protein.<p><a href='/help/pathology_and_biotech_section' target='_top'>More...</a></p>Pathology & Biotechi

<p>This subsection of the 'Pathology and Biotech' section describes the in vivo effects caused by ablation of the gene (or one or more transcripts) coding for the protein described in the entry. This includes gene knockout and knockdown, provided experiments have been performed in the context of a whole organism or a specific tissue, and not at the single-cell level.<p><a href='/help/disruption_phenotype' target='_top'>More...</a></p>Disruption phenotypei

Gls2 and alg6 double mutant cells that transfer Man9GlcNAc2 and that are unable to remove the glucose units as they lack gls2, grow at 37 degrees Celsius, but are characterized by swollen cells, displaying pear, lemon and round shapes and have, when grown at 28 degrees Celsius, a phenotype of growth and morphology almost identical to that of wild-type cells, indicating that facilitation of glycoprotein folding mediated by the interaction of calnexin and monoglucosylated oligosaccharides does not necessarily require cycles of reglucosylation and deglucosylation. The gls2 and alg6 double mutant cells grown in the absence of exogenous stress show an induction of the binding protein (BiP)-encoding mRNA. An incubation of intact gls2 and alg6 mutant cells with [(14)C]glucose for 15 minutes in the absence of 1-deoxynojirimycin (DNJ) leads to the production of protein-linked Glc1Man9GlcNAc2, Man9GlcNAc2 and Man8GlcNAc2.2 Publications

<p>This section describes post-translational modifications (PTMs) and/or processing events.<p><a href='/help/ptm_processing_section' target='_top'>More...</a></p>PTM / Processingi

Molecule processing

Feature keyPosition(s)DescriptionActionsGraphical viewLength
<p>This subsection of the 'PTM / Processing' section denotes the presence of an N-terminal signal peptide.<p><a href='/help/signal' target='_top'>More...</a></p>Signal peptidei1 – 25Sequence analysisAdd BLAST25
<p>This subsection of the 'PTM / Processing' section describes the extent of a polypeptide chain in the mature protein following processing or proteolytic cleavage.<p><a href='/help/chain' target='_top'>More...</a></p>ChainiPRO_000037278226 – 923Glucosidase 2 subunit alphaAdd BLAST898

Amino acid modifications

Feature keyPosition(s)DescriptionActionsGraphical viewLength
<p>This subsection of the <a href="http://www.uniprot.org/help/ptm%5Fprocessing%5Fsection">PTM / Processing</a> section specifies the position and type of each covalently attached glycan group (mono-, di-, or polysaccharide).<p><a href='/help/carbohyd' target='_top'>More...</a></p>Glycosylationi262N-linked (GlcNAc...) asparagineSequence analysis1
Glycosylationi563N-linked (GlcNAc...) asparagineSequence analysis1
Glycosylationi822N-linked (GlcNAc...) asparagineSequence analysis1

Keywords - PTMi

Glycoprotein

Proteomic databases

MaxQB - The MaxQuant DataBase

More...
MaxQBi
Q9US55

PaxDb, a database of protein abundance averages across all three domains of life

More...
PaxDbi
Q9US55

PRoteomics IDEntifications database

More...
PRIDEi
Q9US55

<p>This section provides information on the quaternary structure of a protein and on interaction(s) with other proteins or protein complexes.<p><a href='/help/interaction_section' target='_top'>More...</a></p>Interactioni

<p>This subsection of the <a href="http://www.uniprot.org/help/interaction%5Fsection">'Interaction'</a> section provides information about the protein quaternary structure and interaction(s) with other proteins or protein complexes (with the exception of physiological receptor-ligand interactions which are annotated in the <a href="http://www.uniprot.org/help/function%5Fsection">'Function'</a> section).<p><a href='/help/subunit_structure' target='_top'>More...</a></p>Subunit structurei

Heterodimer of a catalytic subunit alpha (gls2) and a subunit beta (gtb1).

By similarity

Protein-protein interaction databases

The Biological General Repository for Interaction Datasets (BioGRID)

More...
BioGRIDi
279705, 27 interactors

STRING: functional protein association networks

More...
STRINGi
4896.SPAC1002.03c.1

<p>This section provides information on the tertiary and secondary structure of a protein.<p><a href='/help/structure_section' target='_top'>More...</a></p>Structurei

3D structure databases

SWISS-MODEL Repository - a database of annotated 3D protein structure models

More...
SMRi
Q9US55

Database of comparative protein structure models

More...
ModBasei
Search...

<p>This section provides information on sequence similarities with other proteins and the domain(s) present in a protein.<p><a href='/help/family_and_domains_section' target='_top'>More...</a></p>Family & Domainsi

<p>This subsection of the 'Family and domains' section provides information about the sequence similarity with other proteins.<p><a href='/help/sequence_similarities' target='_top'>More...</a></p>Sequence similaritiesi

Belongs to the glycosyl hydrolase 31 family.Sequence analysis

Keywords - Domaini

Signal

Phylogenomic databases

evolutionary genealogy of genes: Non-supervised Orthologous Groups

More...
eggNOGi
KOG1066, Eukaryota

The HOGENOM Database of Homologous Genes from Fully Sequenced Organisms

More...
HOGENOMi
CLU_000631_7_0_1

InParanoid: Eukaryotic Ortholog Groups

More...
InParanoidi
Q9US55

KEGG Orthology (KO)

More...
KOi
K05546

Identification of Orthologs from Complete Genome Data

More...
OMAi
NRTWSIA

Database for complete collections of gene phylogenies

More...
PhylomeDBi
Q9US55

Family and domain databases

Gene3D Structural and Functional Annotation of Protein Families

More...
Gene3Di
2.60.40.1180, 2 hits

Integrated resource of protein families, domains and functional sites

More...
InterProi
View protein in InterPro
IPR033403, DUF5110
IPR011013, Gal_mutarotase_sf_dom
IPR000322, Glyco_hydro_31
IPR025887, Glyco_hydro_31_N_dom
IPR013780, Glyco_hydro_b
IPR017853, Glycoside_hydrolase_SF

Pfam protein domain database

More...
Pfami
View protein in Pfam
PF17137, DUF5110, 1 hit
PF13802, Gal_mutarotas_2, 1 hit
PF01055, Glyco_hydro_31, 1 hit

Superfamily database of structural and functional annotation

More...
SUPFAMi
SSF51445, SSF51445, 1 hit
SSF74650, SSF74650, 1 hit

<p>This section displays by default the canonical protein sequence and upon request all isoforms described in the entry. It also includes information pertinent to the sequence(s), including <a href="http://www.uniprot.org/help/sequence%5Flength">length</a> and <a href="http://www.uniprot.org/help/sequences">molecular weight</a>. The information is filed in different subsections. The current subsections and their content are listed below:<p><a href='/help/sequences_section' target='_top'>More...</a></p>Sequencei

<p>This subsection of the <a href="http://www.uniprot.org/help/sequences%5Fsection">Sequence</a> section indicates if the <a href="http://www.uniprot.org/help/canonical%5Fand%5Fisoforms">canonical sequence</a> displayed by default in the entry is complete or not.<p><a href='/help/sequence_status' target='_top'>More...</a></p>Sequence statusi: Complete.

<p>This subsection of the <a href="http://www.uniprot.org/help/sequences%5Fsection">Sequence</a> section indicates if the <a href="http://www.uniprot.org/help/canonical%5Fand%5Fisoforms">canonical sequence</a> displayed by default in the entry is in its mature form or if it represents the precursor.<p><a href='/help/sequence_processing' target='_top'>More...</a></p>Sequence processingi: The displayed sequence is further processed into a mature form.

Q9US55-1 [UniParc]FASTAAdd to basket
« Hide
        10         20         30         40         50
MRYHGICWFI FQAAIIFAIF GSCQGAFRHQ FKTAEQDGFA RRNRDLAKFQ
60 70 80 90 100
KENLNWNGLF QLNSISYNSG VVSGVFEQQS ENGENQHLFP FSISFLKNDV
110 120 130 140 150
VRFQMDEKSR LEGTVEYEKN ILTKRRFDAS TELGFNERAE VYGKDAHLLE
160 170 180 190 200
QTSTSLTIRY GSHGRFTVIV TFSPFKVEFQ RDGEPQVVLN ERHLLNMEYY
210 220 230 240 250
RPKSSRTPEQ EANGMWDETF DNFHDSKPKG PESVGLDIKF VDYGNVYGVP
260 270 280 290 300
EHTSSLSLKE TNNSDAGYTE PYRLYNVDLF EYEVDSPMSQ YGAIPFMQAH
310 320 330 340 350
KPNSDVAVFW SNAAATWIDV EKESGPSPHS QSTSTHWYSE SGTLDLFIFL
360 370 380 390 400
GPKASDVYES YSALVGRPLL PPLFSIGYHQ CRWNYVSEED VLNVDAKFDE
410 420 430 440 450
VDMPYDTIWL DIEYASKRRY FTWDKATFPN PKAMLEKLDS KSRKLIVILD
460 470 480 490 500
PHIKNDPNYF VSKELIDYNY AVKDKSGVDN YNADCWPGNS VWVDFFNPEA
510 520 530 540 550
QAWWGSLYEF DRFESDKNLW IWNDMNEPSV FRGPETSMHR DAIHYGGWEH
560 570 580 590 600
RDIHNIYGHK CINGTYNGLI KRGEGAVRPF ILTRSFFAGT SALAANWIGD
610 620 630 640 650
TMTTWEHLRG SIPTVLTNGI SGMAFSGADV AGFFGNPDAE LFVRWYETAI
660 670 680 690 700
FYPFFRAHAH IDTKRREPWL YGEPYTSLVR ELLRIRYRLL PTWYTAFYNS
710 720 730 740 750
HTHGFPILYP QFLMHPEDEE GFAIDDQFYV GDSGLLVKPV THPSIDKITI
760 770 780 790 800
YLADDEVYFD LHDHTEYAGK GHQVVPAPLG RVPVLLRGGN ILITRERIRR
810 820 830 840 850
AAELTRNDPF TLTIAVSKIG KNASGFLYLD DGVTFNYKKG EYLIRHFSYE
860 870 880 890 900
NGILTMKDSH SNPPVSPKYS SSQKHLKVER INIYGEQTRK SIKIRKIIDS
910 920
EVTEWDVSVD DSGCIRNPQL FLV
Length:923
Mass (Da):106,282
Last modified:May 1, 2000 - v1
<p>The checksum is a form of redundancy check that is calculated from the sequence. It is useful for tracking sequence updates.</p> <p>It should be noted that while, in theory, two different sequences could have the same checksum value, the likelihood that this would happen is extremely low.</p> <p>However UniProtKB may contain entries with identical sequences in case of multiple genes (paralogs).</p> <p>The checksum is computed as the sequence 64-bit Cyclic Redundancy Check value (CRC64) using the generator polynomial: x<sup>64</sup> + x<sup>4</sup> + x<sup>3</sup> + x + 1. The algorithm is described in the ISO 3309 standard. </p> <p class="publication">Press W.H., Flannery B.P., Teukolsky S.A. and Vetterling W.T.<br /> <strong>Cyclic redundancy and other checksums</strong><br /> <a href="http://www.nrbook.com/b/bookcpdf.php">Numerical recipes in C 2nd ed., pp896-902, Cambridge University Press (1993)</a>)</p> Checksum:iC2377CCD961B4AAF
GO

Sequence databases

Select the link destinations:

EMBL nucleotide sequence database

More...
EMBLi

GenBank nucleotide sequence database

More...
GenBanki

DNA Data Bank of Japan; a nucleotide sequence database

More...
DDBJi
Links Updated
CU329670 Genomic DNA Translation: CAB65603.1

NCBI Reference Sequences

More...
RefSeqi
NP_593490.1, NM_001018924.2

Genome annotation databases

Ensembl fungal genome annotation project

More...
EnsemblFungii
SPAC1002.03c.1; SPAC1002.03c.1:pep; SPAC1002.03c

Database of genes from NCBI RefSeq genomes

More...
GeneIDi
2543277

KEGG: Kyoto Encyclopedia of Genes and Genomes

More...
KEGGi
spo:SPAC1002.03c

<p>This section provides links to proteins that are similar to the protein sequence(s) described in this entry at different levels of sequence identity thresholds (100%, 90% and 50%) based on their membership in UniProt Reference Clusters (<a href="http://www.uniprot.org/help/uniref">UniRef</a>).<p><a href='/help/similar_proteins_section' target='_top'>More...</a></p>Similar proteinsi

<p>This section is used to point to information related to entries and found in data collections other than UniProtKB.<p><a href='/help/cross_references_section' target='_top'>More...</a></p>Cross-referencesi

Sequence databases

Select the link destinations:
EMBLi
GenBanki
DDBJi
Links Updated
CU329670 Genomic DNA Translation: CAB65603.1
RefSeqiNP_593490.1, NM_001018924.2

3D structure databases

SMRiQ9US55
ModBaseiSearch...

Protein-protein interaction databases

BioGRIDi279705, 27 interactors
STRINGi4896.SPAC1002.03c.1

Protein family/group databases

CAZyiGH31, Glycoside Hydrolase Family 31

Proteomic databases

MaxQBiQ9US55
PaxDbiQ9US55
PRIDEiQ9US55

Genome annotation databases

EnsemblFungiiSPAC1002.03c.1; SPAC1002.03c.1:pep; SPAC1002.03c
GeneIDi2543277
KEGGispo:SPAC1002.03c

Organism-specific databases

EuPathDBiFungiDB:SPAC1002.03c
PomBaseiSPAC1002.03c, gls2

Phylogenomic databases

eggNOGiKOG1066, Eukaryota
HOGENOMiCLU_000631_7_0_1
InParanoidiQ9US55
KOiK05546
OMAiNRTWSIA
PhylomeDBiQ9US55

Enzyme and pathway databases

UniPathwayiUPA00957

Miscellaneous databases

Protein Ontology

More...
PROi
PR:Q9US55

Family and domain databases

Gene3Di2.60.40.1180, 2 hits
InterProiView protein in InterPro
IPR033403, DUF5110
IPR011013, Gal_mutarotase_sf_dom
IPR000322, Glyco_hydro_31
IPR025887, Glyco_hydro_31_N_dom
IPR013780, Glyco_hydro_b
IPR017853, Glycoside_hydrolase_SF
PfamiView protein in Pfam
PF17137, DUF5110, 1 hit
PF13802, Gal_mutarotas_2, 1 hit
PF01055, Glyco_hydro_31, 1 hit
SUPFAMiSSF51445, SSF51445, 1 hit
SSF74650, SSF74650, 1 hit

ProtoNet; Automatic hierarchical classification of proteins

More...
ProtoNeti
Search...

MobiDB: a database of protein disorder and mobility annotations

More...
MobiDBi
Search...

<p>This section provides general information on the entry.<p><a href='/help/entry_information_section' target='_top'>More...</a></p>Entry informationi

<p>This subsection of the 'Entry information' section provides a mnemonic identifier for a UniProtKB entry, but it is not a stable identifier. Each reviewed entry is assigned a unique entry name upon integration into UniProtKB/Swiss-Prot.<p><a href='/help/entry_name' target='_top'>More...</a></p>Entry nameiGLU2A_SCHPO
<p>This subsection of the 'Entry information' section provides one or more accession number(s). These are stable identifiers and should be used to cite UniProtKB entries. Upon integration into UniProtKB, each entry is assigned a unique accession number, which is called 'Primary (citable) accession number'.<p><a href='/help/accession_numbers' target='_top'>More...</a></p>AccessioniPrimary (citable) accession number: Q9US55
<p>This subsection of the 'Entry information' section shows the date of integration of the entry into UniProtKB, the date of the last sequence update and the date of the last annotation modification ('Last modified'). The version number for both the entry and the <a href="http://www.uniprot.org/help/canonical%5Fand%5Fisoforms">canonical sequence</a> are also displayed.<p><a href='/help/entry_history' target='_top'>More...</a></p>Entry historyiIntegrated into UniProtKB/Swiss-Prot: May 5, 2009
Last sequence update: May 1, 2000
Last modified: August 12, 2020
This is version 116 of the entry and version 1 of the sequence. See complete history.
<p>This subsection of the 'Entry information' section indicates whether the entry has been manually annotated and reviewed by UniProtKB curators or not, in other words, if the entry belongs to the Swiss-Prot section of UniProtKB (<strong>reviewed</strong>) or to the computer-annotated TrEMBL section (<strong>unreviewed</strong>).<p><a href='/help/entry_status' target='_top'>More...</a></p>Entry statusiReviewed (UniProtKB/Swiss-Prot)
Annotation programFungal Protein Annotation Program

<p>This section contains any relevant information that doesn't fit in any other defined sections<p><a href='/help/miscellaneous_section' target='_top'>More...</a></p>Miscellaneousi

Keywords - Technical termi

Reference proteome

Documents

  1. Schizosaccharomyces pombe
    Schizosaccharomyces pombe: entries and gene names
  2. Glycosyl hydrolases
    Classification of glycosyl hydrolase families and list of entries
  3. PATHWAY comments
    Index of metabolic and biosynthesis pathways
  4. SIMILARITY comments
    Index of protein domains and families
UniProt is an ELIXIR core data resource
Main funding by: National Institutes of Health

We'd like to inform you that we have updated our Privacy Notice to comply with Europe’s new General Data Protection Regulation (GDPR) that applies since 25 May 2018.

Do not show this banner again