Related Experiment Video

Updated: Jul 18, 2026

Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve
09:13

Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve

Published on: June 14, 2017

Recognition of mitochondrial protein precursor lacking arginine at position -2 by mitochondrial processing peptidase:

K Kojima1, E Yamasaki, S Kitada

  • 1Department of Chemistry, Faculty of Science, Kyushu University, Fukuoka 812-8581, Japan.

Journal of Biochemistry
|September 28, 2001
PubMed

Insights

Mitochondrial processing peptidase (MPP) recognizes protein precursors. The amino acid at position -2 is crucial for precise cleavage, with Ala in pre-P450(SCC) influencing MPP recognition and cleavage rates.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzymology

Background:

  • Mitochondrial processing peptidase (MPP) removes N-terminal presequences from mitochondrial protein precursors.
  • Arginine (Arg) at position -2 is a known critical determinant for MPP recognition in many precursors.

Purpose of the Study:

  • To investigate the role of the amino acid at position -2 in the recognition and cleavage of the bovine cytochrome P450 side-chain cleavage enzyme precursor (pre-P450(SCC)) by MPP.
  • To elucidate the structural requirements at position -2 for precise MPP cleavage.

Main Methods:

  • Site-directed mutagenesis of pre-P450(SCC) to introduce different amino acids (Arg, Gly, Met) at position -2.
  • Analysis of MPP cleavage rates and identification of cleavage sites for wild-type and mutant pre-P450(SCC).

Main Results:

  • Replacing Ala at position -2 of pre-P450(SCC) with Arg increased the cleavage rate.
  • Substitution with Gly reduced cleavage rate and introduced an alternative cleavage site.
  • A Met at position -2 maintained cleavage efficiency similar to the wild type.
  • The amino acid at position -2 requires a straight methylene chain for optimal interaction with the MPP S(2) site.

Conclusions:

  • Alanine at position -2 of pre-P450(SCC) acts as a determinant for precise MPP cleavage.
  • The amino acid at position -2 plays a key role in MPP substrate recognition, similar to precursors with Arg at -2.
  • MPP recognition mechanism involves specific interactions at the -2 position, requiring a straight methylene chain.

More Related Videos

Assessment of Submitochondrial Protein Localization in Budding Yeast Saccharomyces cerevisiae08:55

Assessment of Submitochondrial Protein Localization in Budding Yeast Saccharomyces cerevisiae

Published on: July 19, 2021

Mitochondrial Transformation in Baker's Yeast to Study Translation and Respiratory Complex Assembly09:53

Mitochondrial Transformation in Baker's Yeast to Study Translation and Respiratory Complex Assembly

Published on: June 7, 2024

Related Concept Videos

Mitochondrial Protein Sorting01:39

Mitochondrial Protein Sorting

Mitochondria are double-membrane organelles of the eukaryotes involved in cellular metabolism, signaling, ATP synthesis, and programmed cell death.  Each of these processes requires specific proteins and enzymes that must be correctly sorted to the right mitochondrial subcompartment for the proper functioning of the organelle.
Most of these mitochondrial proteins are encoded by the nucleus and imported to the mitochondria as unfolded or loosely folded precursors. Mitochondrial precursors...
Mitochondrial Precursor Proteins01:39

Mitochondrial Precursor Proteins

Mitochondrial precursors are partially unfolded or loosely folded polypeptide chains. Newly synthesized precursors are inhibited from spontaneously folding into their native conformation by the cytosolic chaperones, heat shock proteins 70 (Hsp70), and mitochondrial import stimulation factors (MSFs). Precursors bound to MSFs are guided to the TOM70-TOM37 receptors, while precursors bound to Hsp70  chaperones are targetted to TOM20-TOM22 receptor complexes.
Most of the mitochondrial precursors...
Translocation of Proteins into the Mitochondria01:19

Translocation of Proteins into the Mitochondria

Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
Porin Insertion in the Outer Mitochondrial Membrane01:12

Porin Insertion in the Outer Mitochondrial Membrane

Porins are beta-barrel proteins translocated to the mitochondrial outer membrane through the TOM complex into the intermembrane space. Porin precursors bind TIM chaperones within the intermembrane space and are guided to the Sorting and Assembly Machinery complex or SAM complex on the outer mitochondrial membrane.
Three models describe the assembly of porins by the SAM complex and their insertion into the outer membrane. Model 1 suggests that porins are assembled outside the SAM channel as the...
Protein Transport into the Inner Mitochondrial Membrane01:34

Protein Transport into the Inner Mitochondrial Membrane

Nuclear encoded mitochondrial precursors are imported to the inner membrane in a multistep process involving two separate translocons, TIM22 and TIM23. TIM23 is a cation-selective pore that remains closed by the N terminal segment of the protein. Negative charges on the TIM23 act as a receptor for the incoming precursor, pulling the positively charged matrix-targeting sequence for peptide insertion and translocation.
Transport of mitochondrial precursors across the TIM23 channel is driven by...
Structure of Porins01:21

Structure of Porins

Mitochondria, chloroplasts, and gram-negative bacteria have transmembrane, beta-barrel proteins called porins to mediate the free diffusion of ions and metabolites across the membrane. Mitochondrial porin precursors contain conserved amino acid sequences called beta signals at their C-terminal. Beta signals have a  motif of PoXGXXHyXHy (Po-Polar, X-Any amino acid, G-Glycine, Hy-LargeHydrophobic), which are crucial for precursor recognition to initiate precursor assembly. Beta-barrel precursors...