#METABOLOMICS WORKBENCH McKenna1011_20241108_213403 DATATRACK_ID:5361 STUDY_ID:ST004051 ANALYSIS_ID:AN006697 PROJECT_ID:PR002541
VERSION             	1
CREATED_ON             	July 21, 2025, 12:45 pm
#PROJECT
PR:PROJECT_TITLE                 	Synergistic action of specialized metabolites from divergent biosynthesis in
PR:PROJECT_TITLE                 	human oral microbiome
PR:PROJECT_SUMMARY               	In this project, we report the discovery of two specialized metabolites from one
PR:PROJECT_SUMMARY               	NRPS/PKS gene cluster which are both required for robust biofilm formation in
PR:PROJECT_SUMMARY               	Streptococcus mutans. Submitted are the raw metabolomics data of wildtype
PR:PROJECT_SUMMARY               	Streptococcus mutans Smu102 and its knockouts.
PR:INSTITUTE                     	University of California, Berkeley
PR:LAST_NAME                     	Yao
PR:FIRST_NAME                    	McKenna
PR:ADDRESS                       	Stanley Hall University Dr Berkeley, CA 94720
PR:EMAIL                         	mckenna_yao@berkeley.edu
PR:PHONE                         	4803266096
PR:FUNDING_SOURCE                	National Institute of Dental & Craniofacial Research of the National Institutes
PR:FUNDING_SOURCE                	of Health under Award Number R01DE032732
PR:PUBLICATIONS                  	Synergistic action of specialized metabolites from divergent biosynthesis in
PR:PUBLICATIONS                  	human oral microbiome
#STUDY
ST:STUDY_TITLE                   	Synergistic action of specialized metabolites from divergent biosynthesis in
ST:STUDY_TITLE                   	human oral microbiome
ST:STUDY_SUMMARY                 	In this project, we report the discovery of three specialized metabolites,
ST:STUDY_SUMMARY                 	MC-584, MC-586, and MC-602 from the mutanoclumpin NRPS/PKS gene cluster from
ST:STUDY_SUMMARY                 	Streptococcus mutans, which MC-584 and MC-586 (major metabolites) are both
ST:STUDY_SUMMARY                 	required for robust biofilm formation in Streptococcus mutans. Submitted are the
ST:STUDY_SUMMARY                 	raw metabolomics data of wildtype Streptococcus mutans Smu102 (that has the
ST:STUDY_SUMMARY                 	three metabolites) and its gene knockouts.
ST:INSTITUTE                     	University of California, Berkeley
ST:LAST_NAME                     	Yao
ST:FIRST_NAME                    	McKenna
ST:ADDRESS                       	Stanley Hall University Dr
ST:EMAIL                         	mckenna_yao@berkeley.edu
ST:PHONE                         	4803266096
#SUBJECT
SU:SUBJECT_TYPE                  	Bacteria
SU:SUBJECT_SPECIES               	Streptococcus mutans
SU:TAXONOMY_ID                   	1309
SU:GENOTYPE_STRAIN               	Smu102
#SUBJECT_SAMPLE_FACTORS:         	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data
SUBJECT_SAMPLE_FACTORS           	-	Smu102 wt	Factor:wt | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=wt.mzML
SUBJECT_SAMPLE_FACTORS           	-	mcgn	Factor:mcgn-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=mcgn.mzML
SUBJECT_SAMPLE_FACTORS           	-	mcgm	Factor:mcgm-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=mcgm.mzML
SUBJECT_SAMPLE_FACTORS           	-	ffs	Factor:ffs-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=ffs.mzML
SUBJECT_SAMPLE_FACTORS           	-	mcgd	Factor:mcgd-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=mcgd.mzML
SUBJECT_SAMPLE_FACTORS           	-	mcgb	Factor:mcgb-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=mcgb.mzML
SUBJECT_SAMPLE_FACTORS           	-	1356	Factor:1356-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=1356.mzML
SUBJECT_SAMPLE_FACTORS           	-	1351	Factor:1351-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=1351.mzML
SUBJECT_SAMPLE_FACTORS           	-	1346	Factor:1346-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=1346.mzML
SUBJECT_SAMPLE_FACTORS           	-	1341	Factor:1341-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=1341.mzML
SUBJECT_SAMPLE_FACTORS           	-	mcgo	Factor:mcgO-knockout | Sample source:bacteria	RAW_FILE_NAME(Raw file name)=mcgO.mzML
#COLLECTION
CO:COLLECTION_SUMMARY            	To carry out comparative metabolomic analysis, strains were grown in 5 mL CDM
CO:COLLECTION_SUMMARY            	for 16 hours. Cultures were extracted with 1 volume of 95:5 ethyl
CO:COLLECTION_SUMMARY            	acetate:methanol. Mixtures were vortexed and centrifuged (16,000×g, 5 min). The
CO:COLLECTION_SUMMARY            	organic layer was pipetted into a new tube and dried under N2, then resuspended
CO:COLLECTION_SUMMARY            	in methanol.
CO:SAMPLE_TYPE                   	Bacterial cells
#TREATMENT
TR:TREATMENT_SUMMARY             	No treatment was performed. Only bacterial growth and extraction for comparative
TR:TREATMENT_SUMMARY             	metabolomics.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	For each experiment, strains were first streaked out on Todd Hewitt Broth (THB)
SP:SAMPLEPREP_SUMMARY            	agar plates and grown for two days at 37°C. For S. mutans, single colonies were
SP:SAMPLEPREP_SUMMARY            	then inoculated into Chemically Defined Medium (CDMB) with 4% (v/v) BD™
SP:SAMPLEPREP_SUMMARY            	Difco™ Brain Heart Infusion at 37°C in a standing incubator (CDMB). Growth of
SP:SAMPLEPREP_SUMMARY            	S. mutans in liquid was carried out at 37°C without shaking for 16hours. Unless
SP:SAMPLEPREP_SUMMARY            	specified, cultures were grown with 1M Dextrose (D-glucose).
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	100 μL cell culture equivalent was analyzed on an Agilent Technologies 6545
CH:CHROMATOGRAPHY_SUMMARY        	Accurate-Mass QTOF LC-MS instrument after separation on an Agilent Eclipse Plus
CH:CHROMATOGRAPHY_SUMMARY        	C18 column (4.6×100 mm) 3.5um pour size. Samples were separated using a linear
CH:CHROMATOGRAPHY_SUMMARY        	gradient of 2-98% acetonitrile (v/v) over 30 minutes in H2O with 0.1% formic
CH:CHROMATOGRAPHY_SUMMARY        	acid (v/v) at a flow rate of 0.5 mL/min.
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Agilent Technologies 6545 Accurate-Mass QTOF LC-MS
CH:COLUMN_NAME                   	ZORBAX RR Eclipse Plus C18 (100 mm x 4.6 mm, 3.5 µm)
CH:SOLVENT_A                     	100% Water
CH:SOLVENT_B                     	100% Acetonitrile
CH:FLOW_GRADIENT                 	0-2 min: 2% B; 2-32 min: 2% B - 98% B; 32-37 min: 98% B; 37-37.1 min: 98% B - 2%
CH:FLOW_GRADIENT                 	B; 37-41 min: 2% B
CH:FLOW_RATE                     	0.5 mL/min
CH:COLUMN_TEMPERATURE            	28°C
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Agilent 6445 Q-TOF
MS:INSTRUMENT_TYPE               	QTOF
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	MS data were collected at a fragmentation voltage of 110 V. MS/MS data were
MS:MS_COMMENTS                   	collected throughout the run method using three fragmentation voltages (10, 30,
MS:MS_COMMENTS                   	and 50 V). Data analysis was performed using MassHunter software and comparative
MS:MS_COMMENTS                   	metabolomics with MS-DIAL69. No feature assignment was performed.
MS:MS_RESULTS_FILE               	ST004051_AN006697_Results.txt	UNITS:Peak height	Has m/z:Yes	Has RT:Yes	RT units:Minutes
#END