Standalone Client¶
This page provides some tips on using AACon command line tool.
Executing AACon¶
Type java -jar <AACon jar file name>
java -jar compbio-conservation-1.1.jar
This will print AACon help.
Input¶
Input to AACon can be either
- The list of aligned FASTA formatted sequences
- The Clustal formatted alignment
FASTA example
>UniRef90_Q5VFX
MEWLKIALVAFSGRGNDVDYPCSILGRVANNDKELVNILRNGFFLLTYRMNFSPLPHSSVTSDKGWGCLVRS
SQMLLAHALWRYSANDCRLDHFRDMDTEDSTPFSLHKMVRAVMKKADVFRPEYWTPSQGCEAIRCCVNNAVD
RKLIPPIRVVVCSQGCLLAREICSNLEFGTVLILAPMRCGASRRMTQMMFFSLEHLLHSSACIGVVGGVPQR
SYYILGTSGQRLLYLDPHCMTQEALVSSHAEKAGVVTVTASLVKSVRWDCVDTSCFLGFLVDSFAEWLELRT
CLEELQRRGMEQLLCVDDGVAVGLVDEEIAGWPSEEDVAE
>UniRef90_Q4VBK1
---------------PDTDEPVWILGACYNVKTKKSELLSDSRLWFTYRKKFSPIGGTGPSSDAGWGCMLRC
GQMILAQALWRWDPEKHQPKEYQRFLDKKDSCYSIHQMAQMGVGE-GKSVGEWYGPNTVAQVLKKL----AL
FDDWNSLSVYVSMDNTVVIEDIKKLCDWRPLLLVIPLRMGINS-INPVYIQALKECFKMPQSCGVLGGKPNL
AYYFIGFIDDELIYLDPHTTQQA-VDTESGSAVDDQSFHCQRPHRMKITSLDPSVALGFFCKSEEDFDSWCD
LVQ-------QELLKKRNLRMFELVEKHPSHWPPF-----
>UniRef90_A7RTH8
---MDAACVTYEGVSHETEEDVWILGKRYNIDMGYLNTDVRSRIWLTYRKNFPKIGGTGPTTDSGWGCMLRC
GQMMLAQALWQWDPENEYMQILEAFLDKKDSLYSIHQIAQMGVSE-GKAVGSWFGPNTVAQVLKKL----SA
FDDWSSLCLHVAMDNTVIIEDISN---WRPLVLFIPLRLGLTEM-NVVYNEPLKACFTFKQSLGIIGGRPNH
ATYFIGYFGNNLVYLDPHTTQQTVN-PDELSRIPDGSFHCVYPCRMNIADVDPSVALGFFCKSEEDFDDLCQ
QIKKIIDGKSRPMFEIAK--------DRPQHWPVLE----
Clustal example
CLUSTAL
QUERY/1-328 MEWLKIALVAFSGRGNDVDYPCSILGRVANNDKELVNILRNGFFLLTYRMNFSPLPHSSV
UniRef90_Q4VBK1/1-294 ---------------PDTDEPVWILGACYNVKTKKSELLSDSRLWFTYRKKFSPIGGTGP
UniRef90_A7RTH8/1-303 ---MDAACVTYEGVSHETEEDVWILGKRYNIDMGYLNTDVRSRIWLTYRKNFPKIGGTGP
QUERY/1-328 TSDKGWGCLVRSSQMLLAHALWRYSANDCRLDHFRDMDTEDSTPFSLHKMVRAVMKKADV
UniRef90_Q4VBK1/1-294 SSDAGWGCMLRCGQMILAQALWRWDPEKHQPKEYQRFLDKKDSCYSIHQMAQMGVGEGKS
UniRef90_A7RTH8/1-303 TTDSGWGCMLRCGQMMLAQALWQWDPENEYMQILEAFLDKKDSLYSIHQIAQMGVSEGKA
QUERY/1-328 FRPEYWTPSQGCEAIRCCVNNAVDRKLIPPIRVVVCSQGCLLAREICSNLEFGTVLILAP
UniRef90_Q4VBK1/1-294 VG-EWYGPN----TVAQVLKKLALFDDWNSLSVYVSMDNTVVIEDIKKLCDWRPLLLVIP
UniRef90_A7RTH8/1-303 VG-SWFGPN----TVAQVLKKLSAFDDWSSLCLHVAMDNTVIIEDIS---NWRPLVLFIP
QUERY/1-328 MRCGASRRMTQMMFFSLEHLLHSSACIGVVGGVPQRSYYILGTSGQRLLYLDPHCMTQEA
UniRef90_Q4VBK1/1-294 LRMGIN-SINPVYIQALKECFKMPQSCGVLGGKPNLAYYFIGFIDDELIYLDPH-TTQQA
UniRef90_A7RTH8/1-303 LRLGLT-EMNVVYNEPLKACFTFKQSLGIIGGRPNHATYFIGYFGNNLVYLDPH-TTQQT
QUERY/1-328 LVSSHAEKAGVVTVTASLVKSVRWDCVDTSCFLGFLVDSFAEWLELRTCLEELQRRGMEQ
UniRef90_Q4VBK1/1-294 VDTESGSAVDDQSFHCQRPHRMKITSLDPSVALGFFCKSEEDFDSWCDLVQQ-------E
UniRef90_A7RTH8/1-303 VNPDELSRIPDGSFHCVYPCRMNIADVDPSVALGFFCKSEEDFDDLCQQIK--------K
QUERY/1-328 LLCVDDGVAVGLVDEEIAGWPSEEDVAE
UniRef90_Q4VBK1/1-294 LLKKRNLRMFELVEKHPSHWPPF-----
UniRef90_A7RTH8/1-303 IIDGKSRPMFEIAKDRPQHWPVLE----
The gaps in the alignment can be represented by *, -, space character, X and . (a dot). Other, custom gap characters can also be defined.
Output¶
AACon supports two modes of the output the first and the default mode is RESULT_NO_ALIGNMENT. Only the method names and the conservation score for each position in the alignment are presented. The second output format is RESULT_WITH_ALIGNMEN, where the input, in the form of FASTA formatted alignment, is included at the beginning of the file, followed by the same output as RESULT_NO_ALIGNMENT.
RESULT_NO_ALIGNMENT (and normalised) example is below:
#KABAT 0.7500 0.7500 0.7500 0.3750 0.3750 0.3750 0.9375 0.3750 0.9375 0.3750 0.3750 0.3750 ...
#JORES 0.0000 0.0000 0.0000 1.0000 1.0000 1.0000 1.0000 1.0000 1.0000 1.0000 1.0000 1.0000 ...
#SCHNEIDER 0.6668 0.6668 0.6668 0.3332 0.3332 0.3332 0.7540 0.3332 0.7540 0.3332 0.3332 0.3332 ...
#SHENKIN 0.7789 0.7789 0.7789 0.4600 0.4600 0.4600 0.8448 0.4600 0.8448 0.4600 0.4600 0.4600 ...
#GERSTEIN 0.6667 0.6667 0.6667 0.3333 0.3333 0.3333 0.7540 0.3333 0.7540 0.3333 0.3333 0.3333 ...
#TAYLOR_GAPS 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 0.0000 ...
#TAYLOR_NO_GAPS 0.0000 1.0000 0.5263 0.0000 0.8947 0.0000 0.9474 0.0000 0.0000 0.7895 0.0000 0.8421 ...
#ZVELIBIL 0.1000 0.3000 0.4000 0.1000 0.2000 0.1000 0.3000 0.1000 0.3000 0.2000 0.2000 0.1000 ...
#KARLIN 0.3377 0.3377 0.3377 0.4333 0.3099 0.2641 0.5584 0.2877 0.5584 0.3377 0.4308 0.3377 ...
#ARMON 0.6171 0.6171 0.6171 0.2080 0.1034 0.0625 0.6171 0.1289 0.6171 0.1768 0.2036 0.1027 ...
#THOMPSON 0.7195 0.7269 0.7076 0.4506 0.3942 0.3841 0.4710 0.3744 0.4896 0.4283 0.4041 0.3928 ...
#NOT_LANCET 0.0108 0.0054 0.0538 0.1720 0.1398 0.1075 0.1935 0.1452 0.1935 0.1344 0.2097 0.1344 ...
#MIRNY 0.6667 0.6667 0.6667 0.7540 0.3333 0.7540 0.7540 0.7540 0.7540 0.3333 0.7540 0.3333 ...
#WILLIAMSON 0.8844 0.9531 0.9712 0.5029 0.5109 0.4447 0.6453 0.5525 0.5029 0.5220 0.6764 0.5195 ...
#LANDGRAF 0.7531 0.7268 0.8473 0.5329 0.4288 0.2384 0.4247 0.4698 0.5405 0.2981 0.6343 0.3350 ...
#SANDER 0.0000 0.0000 0.0000 0.1600 0.1067 0.0933 0.1333 0.0267 0.1600 0.1200 0.2000 0.1067 ...
#VALDAR 0.0000 0.0000 0.0000 0.1867 0.1145 0.1145 0.2530 0.0482 0.2530 0.1627 0.2349 0.0904 ...
#SMERFS 0.6250 0.6250 0.6250 0.6250 0.4462 0.3005 0.2324 0.2324 0.2324 0.2324 0.2324 0.2324 ...
RESULT_WITH_ALIGNMENT output example is below:
>QUERY
MEWLKIALVAFSGRGNDVDYPCSILGRVANNDKELVNILRNGFFLLTYRMNFSPLPHSSVTSDKGWGCLVRSSQMLLAHA
LWRYSANDCRLDHFRDMDTEDSTPFSLHKMVRAVMKKADVFRPEYWTPSQGCEAIRCCVNNAVDRKLIPPIRVVVCSQGC
LLAREICSNLEFGTVLILAPMRCGASRRMTQMMFFSLEHLLHSSACIGVVGGVPQRSYYILGTSGQRLLYLDPHCMTQEA
LVSSHAEKAGVVTVTASLVKSVRWDCVDTSCFLGFLVDSFAEWLELRTCLEELQRRGMEQLLCVDDGVAVGLVDEEIAGW
PSEEDVAE
>UniRef90_Q4VBK1
---------------PDTDEPVWILGACYNVKTKKSELLSDSRLWFTYRKKFSPIGGTGPSSDAGWGCMLRCGQMILAQA
LWRWDPEKHQPKEYQRFLDKKDSCYSIHQMAQMGVGE-GKSVGEWYGPNTVAQVLKKL----ALFDDWNSLSVYVSMDNT
VVIEDIKKLCDWRPLLLVIPLRMGINS-INPVYIQALKECFKMPQSCGVLGGKPNLAYYFIGFIDDELIYLDPHTTQQA-
VDTESGSAVDDQSFHCQRPHRMKITSLDPSVALGFFCKSEEDFDSWCDLVQ-------QELLKKRNLRMFELVEKHPSHW
PPF-----
>UniRef90_A7RTH8
---MDAACVTYEGVSHETEEDVWILGKRYNIDMGYLNTDVRSRIWLTYRKNFPKIGGTGPTTDSGWGCMLRCGQMMLAQA
LWQWDPENEYMQILEAFLDKKDSLYSIHQIAQMGVSE-GKAVGSWFGPNTVAQVLKKL----SAFDDWSSLCLHVAMDNT
VIIEDISN---WRPLVLFIPLRLGLTEM-NVVYNEPLKACFTFKQSLGIIGGRPNHATYFIGYFGNNLVYLDPHTTQQTV
N-PDELSRIPDGSFHCVYPCRMNIADVDPSVALGFFCKSEEDFDDLCQQIKKIIDGKSRPMFEIAK--------DRPQHW
PVLE----
#KABAT 3.0000 3.0000 3.0000 6.0000 6.0000 6.0000 1.5000 6.0000 1.5000 6.0000 6.0000 6.0000 ...
(...)
#SMERFS 0.2593 0.2593 0.2593 0.2593 -0.0938 -0.3816 -0.5161 -0.5161 -0.5161 -0.5161 -0.5161 ...
Warning
Please note that only a small part of the output is presented in all examples that follow for the sake of simplicity. Therefore the results are for illustration only. Normally each position of the alignment is given a conservation score by each calculation method.
-f option can be used to specify the output format. For example the command
java -jar compbio-conservation-1.1.jar -i=data.align -m=KABAT -f=RESULT_WITH_ALIGNMENT
outputs the input alignment into the result file.
Calculating conservation¶
Calculate conservation using KABAT conservation method
The alignment is read from the data.align file.
java -jar compbio-conservation-1.1.jar -i=data.align -m=KABAT
The above command will print the results to the console. You should see something like this:
#KABAT 3.0000 3.0000 3.0000 6.0000 6.0000 6.0000 1.5000 6.0000 1.5000 6.0000 6.0000 6.0000 ...
Calculate conservation using two or more methods, for example SANDER and KABAT
Just specify the list of comma separated methods after -m switch like this:
java -jar compbio-conservation-1.1.jar -i=data.align -m=SANDER,KABAT
The results will look something like this:
#KABAT 3.0000 3.0000 3.0000 6.0000 6.0000 6.0000 1.5000 6.0000 1.5000 6.0000 6.0000 6.0000 ...
#SANDER -72.0000 -72.0000 -72.0000 -36.0000 -48.0000 -51.0000 -42.0000 -66.0000 -36.0000 -45.0000 -27.0000 -48.0000 ...
Calculate conservation using all supported conservation methods and make results comparable
java -jar compbio-conservation-1.1.jar -i=data.align -n
The results will be printed to the console. Where -n normalizes all results.
Custom gap character¶
Assuming that the gaps in the alignment are represented by ‘-‘, ‘_’ and ‘x’ symbols the following command will interpret them correctly.
java -jar compbio-conservation-1.1.jar -i=data.align -m=KABAT -g=-,_,x
Running SMERFS with custom parameters¶
Unlike other methods, SMERFS supports a few custom parameters, in particular
- The window width - an integer and an odd number
- Two methods of window scores to columns allocation:
MID_SCORE - gives the window score to the middle column MAX_SCORE - gives the column the highest score of all the windows it belongs to
- A gap percentage cutoff - a float greater than 0 and smaller or equal 1
For example:
java -jar compbio-conservation-1.1.jar -i=data.align -m=SMERFS -s=5,MID_SCORE,0.1
Outputing to a file¶
Use -o option to print the results to the file instead of a console. For example
java -jar compbio-conservation-1.1.jar -i=data.align -n -o=outfile.txt
will produce output.txt results file. Nothing will be printed to the console.
Execution details¶
Use -d option to tell AACon to output its execution details.
java -jar compbio-conservation-1.1.jar -i=data.align -n -d=stat.out -f=RESULT_WITH_ALIGNMENT
The output of the previous command, the context of the stat.out file is below
No methods are request assuming all are required.
No output file is provided, writing results to the standard output.
Setting output format to RESULT_WITH_ALIGNMENT
Using 8 CPUs
Start time: 2017/05/23 09:58:13
Alignment loaded in: 68 ms
Alignment has: 3 sequences.
Alignment length is: 328
KARLIN 17 ms
LANDGRAF 17 ms
SANDER 2 ms
VALDAR 4 ms
SMERFS 4 ms
KABAT 3 ms
SHENKIN 3 ms
SCHNEIDER 5 ms
ARMON 3 ms
JORES 6 ms
GERSTEIN 8 ms
THOMPSON 6 ms
NOT_LANCET 4 ms
MIRNY 9 ms
WILLIAMSON 11 ms
TAYLOR_NO_GAPS 20 ms
ZVELIBIL 21 ms
TAYLOR_GAPS 23 ms
Total calculation time: 0 s
End time: 2017/05/23 09:58:13
Results normalization¶
Different conservation algorithms produce vastly different values. To compare them meaningfully one need to bring the results into the same range. AACon bring the results to the range between 0 and 1 if -n switch is used.
Warning
Normalization should be used carefully, especially when working with very ‘gappy’ alignments, where the occupancy of a single amino acid is observed in aligned columns.
Conservation for large alignments¶
To enable AACon to deal with large alignments (thousands of sequences) let the Java Virtual Machine use more memory with the flag -Xmx<AMOUNT_OF_MEMORY> to your command line as follows:
java -Xmx1G -jar aacon.jar <options>
Where 1G = 1 gigabyte of memory, the same result can be achieved with the following instruction -Xmx1000M. However, floating point numbers like -Xmx1.5G are not allowed.