Identification of a Residue in Helix 2 of Rice Plasma Membrane Intrinsic Proteins That Influences Water Permeability

Abstract

Molecular selection, ion exclusion, and water permeation are well known regulatory mechanisms in aquaporin. Water permeability was found to be diverse in different subgroups of plasma membrane intrinsic proteins (PIPs), even though the residues surrounding the water holes remained the same across the subgroups. Upon homology modeling and structural comparison, a conserved Ala/Ile(Val) residue difference was identified in helix 2 that affected the conformation of the NPA region and consequently influenced the water permeability. The residue difference was found to be conservative within the two subgroups of PIPs in rice as well as in other plants. Func 52% reduction of water transportation. Based on structural analyses and molecular dynamics simulations, we proposed that the difference in water permeability was attributed to the orientation variations of helix 2 that modified waterwater and water-protein interactions. Aquaporins (AQPs)7 are small integral membrane proteins that facilitate water transport across the membranes and are widely distributed in animals, plants, and microbes. It is unique in plants that AQPs form a large family with 35 and 33 members in Arabidopsis and rice, respectively. According to the amino acid sequence, the AQPs could be classified into four subgroups, i.e. the plasma membrane intrinsic proteins (PIPs), the tonoplast intrinsic proteins, the NOD26-like intrinsic protein, and the small basic intrinsic proteins (reviewed in Maurel et al.(1) and references cited therein). Plant PIPs are divided into two phylogenic subgroups, PIP1 and PIP2 (2). The AQP monomer consists of six transmembrane ␣-helices tilted along the plane of the membrane and connected by five loops (A-E). Loop B and D as well as the N and C termini are cytoplasmic (3). Two highly conserved constrictions within the pore of AQPs were proposed by structural analysis of AQP proteins. One is the central constriction formed by two Asn-Pro-Ala (NPA) motifs located on two short ␣-helices and the other is the outer constriction or aromatic/arginine (ar/R) constriction formed by spatial arrangement of four aromatic amino acids. Recently it was found that in addition to facilitating the transport of water, AQPs also can transport other small solutes. Size exclusion at the two main constrictions is one of the mechanisms for water transport and substrate selectivity by AQPs (3, 4). Extensive investigation of the plant PIPs revealed that PIP2 paralogues induced water permeation when expressing on Xenopus oocyte or yeast membranes, whereas PIP1 paralogues were almost inactive (5-10). The sequence difference between the two subgroups, however, is quite subtle, as they share the same residues in main pore constrictions and almost the same hydrophobic residues around the aqueous pathway (6, 11).The first attempt to elucidate the mechanism of distinct water transport activities between the two subgroups was performed in radish PIPs (12). Based on the sequence difference, the authors identified a residue that discriminates PIP1s and PIP2s....

Identification and Expression Analysis of the Barley (Hordeum vulgare L.) Aquaporin Gene Family

“…All HvPIP1s possessed Ile while the HvPIP2s had Val ( S2 Fig ), suggesting the HvPIP1s may have lower water permeability. Further, the difference between OsPIP1s (lower water permeability) and OsPIP2s (higher water permeability) was attributed to a residue in TM2, where OsPIP1s have Ala and OsPIP2s have Ile/Val [ 25 ]. At this position, HvPIP1;1 and HvPIP1;5 had Ala, but all HvPIP2s as well as HvPIP1;2, HvPIP1;3, HvPIP1;4 had Val ( S2 Fig ), suggesting the latter group may be permeable to water and supporting the observation of Besse et al .…”

Single Mutations in the Transmembrane Domains of Maize Plasma Membrane Aquaporins Affect the Activity of Monomers within a Heterotetramer

“…Point mutations, such as those are able to turn an inactive PIP1 into an active channel without any co-expression, have only been reported for rice PIP1;1 and PIP1;3 (Zhang et al, 2010). In that work, the authors identified by comparative modeling and mutagenesis of one residue (Ala102) in TM2 of Os PIP1;3 located at the interface between monomers that induced, upon mutation into Val, a change in the orientation of Ile101, widening constriction region within the pore.…”

Identification and Expression of Nine Oak Aquaporin Genes in the Primary Root Axis of Two Oak Species, Quercus petraea and Quercus robur

“…PIP1s had an Ala residue, whereas PIP2s had a Val or Ile. In Oryza sativa , this residue was found to be involved in the osmotic water permeability of AQPs [48]. The presence of a Val or Ile residue in helix 2 conferred a high permeability to water of PIP2 members, compared to PIP1 members exhibiting a Ala residue at this location.…”