Effectors of Rab GTPases: Rab Binding Specificity and Their Role in Coordination of Rab Function and Localization - Ras Superfamily Small G Proteins: Biology and Mechanisms 2 - page 54

Table 3.3 (continued)
Rab Effector K d Meth. a Reference
Rab25 FIP2 650 nM ITC Lall et al. ( 2013 )
Rab25 MyoVb 537 nM SPR Pylypenko
et al. ( 2013 )
Rab27a Slp2-a 13.4 nM SPR Fukuda ( 2006 )
Rab27a Slp4-a 19.2 nM SPR Fukuda ( 2006 )
Rab27a Slac2-a/melanophilin 112 nM SPR Fukuda ( 2006 )
a Methods used to determine the K d values: FS fluorescence spectroscopy, SPR surface plasmon
resonance, ITC isothermal titration calorimetry.
specific in their interactions and bind to several Rab proteins. The impact of this
will be discussed in the following chapter.
3.4 Promiscuity of Rab Effector Proteins
A systematic screening for novel Rab effectors based on the yeast two-hybrid assay
performed by the Fukuda group revealed that many effectors exhibit, at least
in vitro, a broad Rab binding specificity (Fukuda et al. 2008 ). In most cases, such
effectors contain two or more Rab binding sites (see below). There are however a
few examples of effectors that are able to bind many Rab proteins through the same
domain. The best known are the two functionally related inositol 5-phosphatases
OCRL and INPP5B. Most studies focused on OCRL. OCRL can interact, at least
in vitro, with a subset of sixteen Rab GTPases (including isoforms), i.e., Rab1,
Rab3, Rab5, Rab6, Rab8, Rab13, Rab14, Rab22b, and Rab35 (Hyvola et al. 2006 ;
Fukuda et al. 2008 ). The Rab binding site of OCRL has been mapped to a region
between the phosphatase and the Rho-GAP domains (the ASH domain) (Hyvola
et al. 2006 ). It consists of a relatively short domain of about 130 amino acids.
Possible explanations for the binding of such a large variety of different Rab
proteins can be deduced from the structure of the Rab binding domain of OCRL
in complex with Rab8a (Hou et al. 2011 ). This study shows that the hydrophobic
triad appears to have only a minor role in OCRL binding, which could be one reason
for the capacity of OCRL to interact with many Rabs. Furthermore, the interaction
between OCRL and the Rab protein occurs mostly via polar interactions, and most
of the intermolecular hydrogen bonds formed involve the main chain atoms of the
Rab molecule and side chains that are conserved between different Rab proteins.
This interaction is less sequence sensitive compared to other Rab:effector interac-
tions, and may explain the promiscuity of OCRL in Rab recognition.
Another example of promiscuity in Rab binding is the Rab5 effector
Rabenosyn-5. Rabenosyn-5 has distinct Rab4 and Rab5 binding sites (see below);
however the minimal Rab4 binding domain also binds with similar affinity to
Rab14 (subset-1) (Eathiraj et al. 2005 ). In addition, the Rab5 binding domain