Signature motif identification and enzymatic characterization of a protein tyrosine phosphatase in <i>Metarhizium anisopliae</i>.

Ze TAN; Pei ZHU; Zhenlun LI; Shuiying YANG

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Signature motif identification and enzymatic characterization of a protein tyrosine phosphatase in Metarhizium anisopliae.

Author: Ze TAN ¹ ; Pei ZHU ² ; Zhenlun LI ² ; Shuiying YANG ¹
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1. College of Plant Protection, Southwest University, Chongqing 400715, China.
2. College of Resources and Environment, Southwest University, Chongqing 400715, China.
Publication Type:Journal Article
Keywords: Metarhizium anisopliae; gene expression; phosphatase characteristic; protein tyrosine phosphatase; signature motif of protein
MeSH: Metarhizium/genetics*; Protein Tyrosine Phosphatases/chemistry*; Amino Acid Motifs; Recombinant Proteins/biosynthesis*; Amino Acid Sequence; Pichia/metabolism*; Fungal Proteins/chemistry*; Substrate Specificity; Saccharomycetales
From: Chinese Journal of Biotechnology 2025;41(9):3579-3588
CountryChina
Language:Chinese
Abstract: Protein tyrosine phosphatases (PTPs, EC 3.1.3.48) are key regulators of cellular processes, with the catalytic activity attributed to the conserved motif (H/V)CX₅R(S/T), where cysteine and arginine residues are critical. Previous studies revealed that alternative splicing of extracellular phosphatase mRNA precursors in Metarhizium anisopliae generated two distinct transcripts, with the longer sequence containing a novel HCPTPMLS motif resembling PTP signatures but lacking the arginine residue. To identify the novel signature motif and characterize its enzymatic properties, we heterologously expressed and purified both proteins in Pichia pastoris and comprehensively characterized their enzymatic properties. The protein containing the HCPTPMLS motif (designated as L-protein) exhibited the highest activity at pH 5.5 and a strong preference for pTyr substrates. Its phosphatase activity was inhibited by Ag⁺, Zn²⁺, Cu²⁺, molybdate, and tungstate, but enhanced by Ca²⁺ and EDTA. AcP101 (lacking HCPTPMLS) showed the maximal activity at pH 6.5 and a strong preference toward pNPP (P < 0.05), with the activity inhibited by NaF and tartrate, but enhanced by Mg²⁺ and Mn²⁺. Functional analysis confirmed that the L-protein retained the PTP activity despite the absence of arginine in its signature motif, while AcP101 functioned as an acid phosphatase. This study provides the first functional validation of an arginine-deficient PTP motif, expanding the definition of PTP signature motifs and offering new insights for phosphatase classification.