VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

VESICLE-INDUCING PROTEIN IN PLASTID1 (VIPP1) is conserved among oxygenic photosynthetic organisms and appears to have diverged from the bacterial PspA protein. VIPP1 localizes to the chloroplast envelope and thylakoid membrane, where it forms homooligomers of high molecular mass. Although multiple roles of VIPP1 have been inferred, including thylakoid membrane formation, envelope maintenance, membrane fusion, and regulation of photosynthetic activity, its precise role in chloroplast membrane quality control remains unknown. VIPP1 forms an oligomer through its amino-terminal domain and triggers membrane fusion in an Mg2+-dependent manner. We previously demonstrated that Arabidopsis (Arabidopsis thaliana) VIPP1 also exhibits dynamic complex disassembly in response to osmotic and heat stresses in vivo. These results suggest that VIPP1 mediates membrane fusion/remodeling in chloroplasts. Considering that protein machines that regulate intracellular membrane fusion/remodeling events often require a capacity for GTP binding and/or hydrolysis, we questioned whether VIPP1 has similar properties. We conducted an in vitro assay using a purified VIPP1-His fusion protein expressed in Escherichia coli cells. VIPP1-His showed GTP hydrolysis activity that was inhibited competitively by an unhydrolyzable GTP analog, GTPγS, and that depends on GTP binding. It is particularly interesting that the ancestral PspA from E. coli also possesses GTP hydrolysis activity. Although VIPP1 does not contain a canonical G domain, the amino-terminal α-helix was found to be important for both GTP binding and GTP hydrolysis as well as for oligomer formation. Collectively, our results reveal that the properties of VIPP1/PspA are similar to those of GTPases.

Original languageEnglish
Pages (from-to)328-338
Number of pages11
JournalPlant Physiology
Volume177
Issue number1
DOIs
Publication statusPublished - May 1 2018

Fingerprint

GTP Phosphohydrolases
guanosinetriphosphatase
Chloroplasts
chloroplasts
Membranes
Guanosine Triphosphate
Proteins
proteins
Membrane Fusion
Hydrolysis
hydrolysis
Thylakoids
Arabidopsis
thylakoids
In Vitro Techniques
Escherichia coli
Osmoregulation
Heat-Shock Response
Intracellular Membranes
bacterial proteins

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Cite this

VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro. / Ohnishi, Norikazu; Zhang, Lingang; Sakamoto, Wataru.

In: Plant Physiology, Vol. 177, No. 1, 01.05.2018, p. 328-338.

Research output: Contribution to journalArticle

@article{e5c8d30911874ac3ae6ad2bb06d43454,
title = "VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro",
abstract = "VESICLE-INDUCING PROTEIN IN PLASTID1 (VIPP1) is conserved among oxygenic photosynthetic organisms and appears to have diverged from the bacterial PspA protein. VIPP1 localizes to the chloroplast envelope and thylakoid membrane, where it forms homooligomers of high molecular mass. Although multiple roles of VIPP1 have been inferred, including thylakoid membrane formation, envelope maintenance, membrane fusion, and regulation of photosynthetic activity, its precise role in chloroplast membrane quality control remains unknown. VIPP1 forms an oligomer through its amino-terminal domain and triggers membrane fusion in an Mg2+-dependent manner. We previously demonstrated that Arabidopsis (Arabidopsis thaliana) VIPP1 also exhibits dynamic complex disassembly in response to osmotic and heat stresses in vivo. These results suggest that VIPP1 mediates membrane fusion/remodeling in chloroplasts. Considering that protein machines that regulate intracellular membrane fusion/remodeling events often require a capacity for GTP binding and/or hydrolysis, we questioned whether VIPP1 has similar properties. We conducted an in vitro assay using a purified VIPP1-His fusion protein expressed in Escherichia coli cells. VIPP1-His showed GTP hydrolysis activity that was inhibited competitively by an unhydrolyzable GTP analog, GTPγS, and that depends on GTP binding. It is particularly interesting that the ancestral PspA from E. coli also possesses GTP hydrolysis activity. Although VIPP1 does not contain a canonical G domain, the amino-terminal α-helix was found to be important for both GTP binding and GTP hydrolysis as well as for oligomer formation. Collectively, our results reveal that the properties of VIPP1/PspA are similar to those of GTPases.",
author = "Norikazu Ohnishi and Lingang Zhang and Wataru Sakamoto",
year = "2018",
month = "5",
day = "1",
doi = "10.1104/pp.18.00145",
language = "English",
volume = "177",
pages = "328--338",
journal = "Plant Physiology",
issn = "0032-0889",
publisher = "American Society of Plant Biologists",
number = "1",

}

TY - JOUR

T1 - VIPP1 Involved in Chloroplast Membrane Integrity Has GTPase Activity in Vitro

AU - Ohnishi, Norikazu

AU - Zhang, Lingang

AU - Sakamoto, Wataru

PY - 2018/5/1

Y1 - 2018/5/1

N2 - VESICLE-INDUCING PROTEIN IN PLASTID1 (VIPP1) is conserved among oxygenic photosynthetic organisms and appears to have diverged from the bacterial PspA protein. VIPP1 localizes to the chloroplast envelope and thylakoid membrane, where it forms homooligomers of high molecular mass. Although multiple roles of VIPP1 have been inferred, including thylakoid membrane formation, envelope maintenance, membrane fusion, and regulation of photosynthetic activity, its precise role in chloroplast membrane quality control remains unknown. VIPP1 forms an oligomer through its amino-terminal domain and triggers membrane fusion in an Mg2+-dependent manner. We previously demonstrated that Arabidopsis (Arabidopsis thaliana) VIPP1 also exhibits dynamic complex disassembly in response to osmotic and heat stresses in vivo. These results suggest that VIPP1 mediates membrane fusion/remodeling in chloroplasts. Considering that protein machines that regulate intracellular membrane fusion/remodeling events often require a capacity for GTP binding and/or hydrolysis, we questioned whether VIPP1 has similar properties. We conducted an in vitro assay using a purified VIPP1-His fusion protein expressed in Escherichia coli cells. VIPP1-His showed GTP hydrolysis activity that was inhibited competitively by an unhydrolyzable GTP analog, GTPγS, and that depends on GTP binding. It is particularly interesting that the ancestral PspA from E. coli also possesses GTP hydrolysis activity. Although VIPP1 does not contain a canonical G domain, the amino-terminal α-helix was found to be important for both GTP binding and GTP hydrolysis as well as for oligomer formation. Collectively, our results reveal that the properties of VIPP1/PspA are similar to those of GTPases.

AB - VESICLE-INDUCING PROTEIN IN PLASTID1 (VIPP1) is conserved among oxygenic photosynthetic organisms and appears to have diverged from the bacterial PspA protein. VIPP1 localizes to the chloroplast envelope and thylakoid membrane, where it forms homooligomers of high molecular mass. Although multiple roles of VIPP1 have been inferred, including thylakoid membrane formation, envelope maintenance, membrane fusion, and regulation of photosynthetic activity, its precise role in chloroplast membrane quality control remains unknown. VIPP1 forms an oligomer through its amino-terminal domain and triggers membrane fusion in an Mg2+-dependent manner. We previously demonstrated that Arabidopsis (Arabidopsis thaliana) VIPP1 also exhibits dynamic complex disassembly in response to osmotic and heat stresses in vivo. These results suggest that VIPP1 mediates membrane fusion/remodeling in chloroplasts. Considering that protein machines that regulate intracellular membrane fusion/remodeling events often require a capacity for GTP binding and/or hydrolysis, we questioned whether VIPP1 has similar properties. We conducted an in vitro assay using a purified VIPP1-His fusion protein expressed in Escherichia coli cells. VIPP1-His showed GTP hydrolysis activity that was inhibited competitively by an unhydrolyzable GTP analog, GTPγS, and that depends on GTP binding. It is particularly interesting that the ancestral PspA from E. coli also possesses GTP hydrolysis activity. Although VIPP1 does not contain a canonical G domain, the amino-terminal α-helix was found to be important for both GTP binding and GTP hydrolysis as well as for oligomer formation. Collectively, our results reveal that the properties of VIPP1/PspA are similar to those of GTPases.

UR - http://www.scopus.com/inward/record.url?scp=85049627140&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85049627140&partnerID=8YFLogxK

U2 - 10.1104/pp.18.00145

DO - 10.1104/pp.18.00145

M3 - Article

C2 - 29622686

AN - SCOPUS:85049627140

VL - 177

SP - 328

EP - 338

JO - Plant Physiology

JF - Plant Physiology

SN - 0032-0889

IS - 1

ER -