METHODS AND COMPOSITIONS FOR PROTEIN SEQUENCING
Aspects of the application provide methods of identifying and sequencing proteins, polypeptides, and amino acids, and compositions useful for the same. In some aspects, the application provides methods of obtaining data during a degradation process of a polypeptide, and outputting a sequence representative of the polypeptide. In some aspects, the application provides amino acid recognition molecules comprising a shielding element that enhances photostability in polypeptide sequencing reactions.
1 . A method comprising:
obtaining data during a degradation process of a polypeptide;
analyzing the data to determine portions of the data corresponding to amino acids that are sequentially exposed at a terminus of the polypeptide during the degradation process; and
outputting an amino acid sequence representative of the polypeptide.
2 . The method of claim 1 , wherein the data is indicative of amino acid identity at the terminus of the polypeptide during the degradation process.
3 . The method of claim 2 , wherein the data is indicative of a signal produced by one or more amino acid recognition molecules binding to different types of terminal amino acids at the terminus during the degradation process.
4 . The method of claim 1 , wherein the data is indicative of a luminescent signal generated during the degradation process.
5 . The method of claim 1 , wherein the data is indicative of an electrical signal generated during the degradation process.
6 . The method of claim 1 , wherein analyzing the data further comprises detecting a series of cleavage events and determining the portions of the data between successive cleavage events.
7 . The method of claim 1 , wherein analyzing the data further comprises determining a type of amino acid for each of the individual portions.
8 . The method of claim 1 , wherein each of the individual portions comprises a pulse pattern, and analyzing the data further comprises determining a type of amino acid for one or more of the portions based on its respective pulse pattern.
9 . The method of claim 8 , wherein determining the type of amino acid further comprises identifying an amount of time within a portion when the data is above a threshold value and comparing the amount of time to a duration of time for the portion.
10 . The method of claim 8 , wherein determining the type of amino acid further comprises identifying at least one pulse duration for each of the one or more portions.
11 . The method of claim 8 , wherein determining the type of amino acid further comprises identifying at least one interpulse duration for each of the one or more portions.
12 . The method of claim 1 , wherein the amino acid sequence includes a series of amino acids corresponding to the portions.
13 . A system comprising:
at least one hardware processor; and
at least one non-transitory computer-readable storage medium storing processor-executable instructions that, when executed by the at least one hardware processor, cause the at least one hardware processor to perform the method of claim 1 .
14 . At least one non-transitory computer-readable storage medium storing processor-executable instructions that, when executed by at least one hardware processor, cause the at least one hardware processor to perform the method of claim 1 .
15 - 283 . (canceled)
284 . The method of claim 10 , wherein the pulse pattern of each of the one or more portions comprises a mean pulse duration of between about 1 millisecond and about 10 seconds.
285 . The method of claim 284 , wherein the mean pulse duration is between about 10 milliseconds and about 100 milliseconds or between about 100 milliseconds and about 500 milliseconds.
286 . The method of claim 10 , wherein the pulse pattern of one type of amino acid is different from the pulse pattern of another type of amino acid by a mean pulse duration of at least 10 milliseconds.
287 . The method of claim 286 , wherein the pulse pattern of one type of amino acid is different from the pulse pattern of another type of amino acid by a mean pulse duration of between about 10 milliseconds and about 100 milliseconds or between about 100 milliseconds and about 10 seconds.