Biomolecular NMR Assignments

, Volume 12, Issue 1, pp 63–68 | Cite as

Backbone and Ile-δ1, Leu, Val methyl 1H, 15N, and 13C, chemical shift assignments for Rhizopus chinensis lipase

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Abstract

Lipase r27RCL is a 296-residue, 33 kDa monomeric enzyme with high ester hydrolysis activity, which has significant applications in the baking, paper and leather industries. The lipase gene proRCL from Rhizopus microsporus var. chinensis (also Rhizopus chinensis) CCTCC M201021 was cloned as a fusion construct C-terminal to a maltose-binding protein (MBP) tag, and expressed as MBP-proRCL in an Escherichia coli BL21 trxB (DE3) expression system with uniform 2H,13C,15N-enrichment and Ile-δ1, Leu, and Val 13CH3 methyl labeling. The fusion protein was hydrolyzed by Kex2 protease at the recognition site Lys-Arg between residues −29 and −28 of the prosequence, producing the enzyme form called r27RCL. Here we report extensive backbone 1H, 15N, and 13C, as well as Ile-δ1, Leu, and Val side chain methyl, NMR resonance assignments for r27RCL.

Keywords

NMR resonance assignments Rhizopus microsporus var. chinensis lipase 

Abbreviations

CSI

Chemical shift index

DSS

2,2-Dimethyl-2-silapentane-5-sulfonic acid

HSQC

Heteronuclear single quantum coherence

TROSY

Transverse relaxation optimized spectroscopy

NOESY

Nuclear overhauser effect spectroscopy

RCL

Rhizopus chinensis lipase

IPTG

Isopropyl β-d-1-thiogalactopyranoside

Notes

Acknowledgements

We thank Profs. J. Hunt and T. Szyperski for helpful discussions on the RCL lipase project. Financial support from the High-end Foreign Experts Recruitment Program (GDW20123200113), Six Talent Peaks Project in Jiangsu Province (NY-010), 333 Project in Jiangsu Province (BRA2015316), NSFC (31671799), and the 111 Project (111-2-06) are greatly appreciated. This work was also supported as a Community Outreach Project of the NIH NIGMS Protein Structure Initiative, Grant U54 GM094597.

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Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of BiotechnologyJiangnan UniversityWuxiChina
  2. 2.State Key Laboratory of Food Science and TechnologyJiangnan UniversityWuxiChina
  3. 3.Center for Advanced Biotechnology and Medicine, Department of Molecular Biology and BiochemistryRutgers, The State University of New JerseyPiscatawayUSA
  4. 4.Department of Biochemistry and Molecular BiologyRobert Wood Johnson Medical School, Rutgers, The State University of New JerseyPiscatawayUSA
  5. 5.Northeast Structural Genomics ConsortiumPiscatawayUSA

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