XYZ4D is a NMRspy extension written in JAVA to assist the assignment of backbone resonances of large proteins without the use of deuterium and specific labeling.
It is based on a strategy recently developed in our lab, which extends the size limit for structure determination by NMR spectroscopy to 42 kDa for monomeric proteins and to 65 kDa for differentially labeled multimeric proteins.
XYZ4D is capable of doing following tasks:
- Manage a set of spectra (2D-TROSY-HSQC, 3D-TROSY-HNCA, 3D-TROSY-HN(CO)CA, 3D-MQ-CCH-TOCSY and 4D-13C,15N-edited NOESY) that are used in the strategy;
- Perform calibration and peak picking;
- Group HC-NH NOE and Cα-NH (HNCA,HN(CO)CA) correlations that have identical NH chemical shifts into clusters;
- Identify spin-systems by separating intra-residue and sequential HC-NH NOE correlations from other inter-residue NOEs observed in the 4D-NOESY spectrum with the use of HNCA and CCH-TOCSY spectra;
- Establish fragments from clusters by matching the intra-residue spin-system of one cluster with the sequential spin-system of another cluster;
- Map the fragments onto the protein sequence.
- Utilizes 2D-TROSY-HSQC, 3D-TROSY-HNCA, 3D-TROSY-HN(CO)CA, 3D-MQ-CCH-TOCSY and 4D-13C,15N-edited NOESY
- Backbone assignments can be obtained with minimal human intervention.
- The reliability of each assigned cluster is indicated using a trustworthy score.
- Graphical interfaces for users to manually check through all the tasks and make corrections should any error occur.
- The whole program is divided into seven modules (Project Preparation, Spectra Calibration, Cluster Identification, CCH & 4DNOE Inspection, Spin-system Identification, Cluster Mapping, Backbone Assignment), which correspond with the manual assignment steps of our new strategy.
- Each module provides a semi-automated approach with a fully automated option
XYZ4D is provided as an extension of NMRspy which can be freely downloaded at this website. Carefully read the instructions on using XYZ4D. Some frequently asked questions are also listed out for your information, together with the workaround. Feel free to contact us if you need further support.
Start NMRspy, click “Extensions” on the menu of its Control Panel. You will find a “XYZ4D” button on the pull-down menu. Click on it to start the XYZ4D extension.