In a mass spectrum of fragment ions obtained by dissociating peptide-derived ions using the technique of irradiating the ions with hydrogen radicals, either pairs of a-type and c-type ions or those of z-type and z-type ions are characteristically observed. Since the mass difference of those ion pairs is previously known, a pair peak searcher 92 searches for pair peaks having a predetermined mass difference in a mass spectrum created by a mass spectrum creator 91, and adds to the detected pair peaks a piece of information indicating that they are pairs of a-type and c-type ions or those of x-type and z-type ions. When estimating the amino acid sequence of the peptide by a database search, a protein identifier 93 uses the ion-pair information in addition to the m/z value of each peak, whereby the accuracy of the estimation or identification the amino acid sequence can be improved.

Methods and systems for determining amino acid sequence of a polypeptide or protein from mass spectrometry data is provided, using a weighted de Bruijn graph. Extracted and purified protein is cleaved into a mixture of peptide and then analyzed using mass spectrometry. A list of peptide sequences is derived from mass spectrometry fragment data by de novo sequencing, and amino acid confidence scores are determined from peak fragment ion intensity. A weighted de Bruijn graph is constructed for the list of peptide sequences having node weights defined by k1 mer confidence scores. At least one contig is assembled from the de Bruijn graph by identifying node weights having the highest k1 mer confidence scores.

The present invention relates to the field of biochemistry, more particularly to proteomics, more particularly to protein sequencing, even more particularly to single molecule peptide sequencing. The invention discloses methods for single molecule protein sequencing and/or amino acid identification using cleavage inducing agents which are not specific for one particular amino acid, cleave polypeptides step by step from the N-terminus onwards and provide information on the identity of the cleaved amino acids based on the kinetics.

An oligopeptide has the amino acid sequence of Thr-Val-Asp-Ser-Cys-Leu-Thr (SEQ ID NO: 1) with addition or insertion of a (poly)peptide to the oligopeptide, or substitution of a (poly)peptide for a part of amino acids of the oligopeptide.

The present disclosure relates to systems and methods for analysis of proteins, more in particular to nanopore systems, devices and methods for single-molecule protein analysis and sequencing. Provided is a method for translocating a target protein through a nanopore, the nanopore being comprised in a membrane separating a fluidic chamber of a nanopore system into a cis side and a trans side, comprising: (a) allowing a protein translocase in solution to capture and form a complex with the target protein to be translocated; (b) contacting the translocase-target protein complex with the cis side of the nanopore and allowing for translocation of the target protein to the trans side; wherein the nanopore system has a cis to trans electro-osmotic force (EOF) resulting from a large net ionic current flow cis-to-trans relative to the total ionic current flow, so that the target protein is captured in the nanopore with on top of the nanopore the translocase controlling the translocation.

The present systems and methods introduce deep learning to de novo peptide sequencing from tandem mass spectrometry data, and in particular mass spectrometry data obtained by data-independent acquisition. The systems and methods achieve improvements in sequencing accuracy over existing systems and methods and enables complete assembly of novel protein sequences without assisting databases. To sequence peptides from mass spectrometry data obtained by data-independent acquisition, precursor profiles representing intensities of one or more precursor ion signals associated with a precursor retention time and fragment ion spectra representing signals from fragment ions and fragment retention times are fed into a neural network.

A bioinspired protein pore-based nanostructure that can provide selective, real-time sampling of protein-protein interactions at single-molecule resolution. This modular nanostructure relied on a single polypeptide chain that encompassed a heavily truncated outer membrane protein, a highly flexible connector, a protein receptor element, as well as a polypeptide adapter. The presence of a protein ligand analyte in solution produced reversible binding and release events, in the form of discrete and stochastic current transitions between open substates of the transmembrane pore, the nature of which depend on both the amount of protein ligand analyte and the strength of the transient PPIs in aqueous phase.

The method for aiding ALS detection provided by the present invention includes determining a profile of signal peptides contained in a bodily fluid from a test subject, and comparing the signal peptide profile thus determined for the test subject with a previously-determined profile of signal peptides in a bodily fluid from a healthy subject. The presence of a difference between the signal peptide profile of the test subject and the signal peptide profile of the healthy subject at a specific molecular weight is then associated with the test subject's suffering from or developing ALS.

Provided herein are compounds of Formulae (I) and (II), which comprise polypeptidyl groups. Also provided herein are methods of preparing compounds of Formulae (I) and (II). Further provided herein are methods of sequencing a polypeptide by reaction of compounds of Formula (II) with peptidases.

Disclosed are thin inorganic membranes having a defined topography that includes pores having a defined diameter of nanometer and sub-nanometer diameter. The thin membranes are resistant to protein denaturing agents, and may be employed in analytical and clinical methods for identifying single amino acid residues within the sequence of a protein, and the pores are other than MspA pores. Methods for making a thin inorganic membrane with nanopore and sub-nanopore topography and conical cone structure are also disclosed. The thin inorganic membrane may be comprised of any denaturant-resistant materials, such as silicon nitride. A method for manufacturing the thin inorganic membrane with nanopores is also provided, and provides a thin surface with a defined conical topography, the nanopores being provided on the membrane surface with an electron beam sputtering technique.