Purification and characterization of a papaya (Carica papaya L.) pectin methylesterase isolated from a commercial papain preparation

Prasanna Vasu; Brett J Savary; Randall G Cameron

doi:10.1016/j.foodchem.2012.01.042

Purification and characterization of a papaya (Carica papaya L.) pectin methylesterase isolated from a commercial papain preparation

Food Chem. 2012 Jul 15;133(2):366-72. doi: 10.1016/j.foodchem.2012.01.042. Epub 2012 Jan 24.

Authors

Prasanna Vasu¹, Brett J Savary², Randall G Cameron³

Affiliations

¹ Arkansas Biosciences Institute, Arkansas State University, Jonesboro, AR, USA.
² Arkansas Biosciences Institute, Arkansas State University, Jonesboro, AR, USA; College of Agriculture and Technology, Arkansas State University, Jonesboro, AR, USA. Electronic address: bsavary@astate.edu.
³ US Horticultural Research Laboratory, Agricultural Research Service, US Department of Agriculture, Ft. Pierce, FL, USA.

PMID: 25683408
DOI: 10.1016/j.foodchem.2012.01.042

Abstract

We purified a Carica papaya pectin methylesterase (CpL-PME; EC 3.1.1.11) from a commercial papain preparation. This CpL-PME was separated from the abundant cysteine endopeptidases activities using sequential hydrophobic interaction and cation-exchange chromatographies and then purified by affinity chromatography using Sepharose-immobilized kiwi PME inhibitor protein to obtain a single electrophoretically homogeneous protein. The enzyme was purified 92-fold with 38% yield, providing a specific activity of 1200 U/mg. The molecular weight was determined to be 35,135 by MALDI-TOF-MS in linear mode. MALDI-TOF-MS peptide mass fingerprinting following trypsin digestion indicated CpL-PME represents a novel Carica PME isoform. The CpL-PME required salt for activity, and it showed a broad activity range (pH 6-9) and moderate thermostability (optimum ca. 70°C). A calcium-insensitive methylated lime pectin treated with CpL-PME to reduce degree of methylesterification by 6% converted the substrate to high calcium sensitivity, indicating a processive mode of action. These properties support further research to apply CpL-PME to tailor pectin nanostructure.

Keywords: Biochemical characterization; Carica papaya; Chromatography purification; Fruit latex; Liquipanol; MALDI-TOF-MS; Pectin methylesterase.

Publication types

Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Carica / chemistry*
Chromatography / methods*
Fruit / chemistry*
Mass Spectrometry
Papain / chemistry*
Pectins / chemistry*

Substances

Pectins
Papain