Crystallization of a proteolyzed form of the horse pancreatic lipase-related protein 2: structural basis for the specific detergent requirement.

The research article focuses on the crystallization process of a proteolyzed form of a horse pancreatic lipase-related protein (PLRP2), specifically in relation to the need for specific detergents like N,N-dimethyldecylamine-beta-oxide (DDAO) for stabilization and crystallization.

The Proteins and Their Characteristics

• The focus of this research article is on horse pancreatic lipase-related proteins (PLRP1 and PLRP2), and they are produced by the pancreas, together with pancreatic lipase (PL).
• These proteins exhibit a common two-domain organization – N-terminal catalytic domain and a C-terminal domain involved in colipase binding, especially in pancreatic lipase.
• Despite the high level of sequence identity, these proteins demonstrate different enzymatic properties.

The Flap Region & Proteolytic Cleavage

• The ‘flap’ region of PLRP2, specifically between Ser245 and Thr246, is inherently sensitive to proteolytic cleavage – the breaking of a bond in a protein by an enzyme.
• It is suggested that this sensitivity might be complicating the crystallization process of the PLRP2.

The Role of Specific Detergent

• The research illustrates how the proteolyzed form of PLRP2 is crystallized only after the specific stabilization of the flap region using a specific detergent – N,N-dimethyldecylamine-beta-oxide (DDAO).
• This stabilization and crystallization process were carried out via the method of hanging-drop vapour-diffusion at 291K and only in the presence of DDAO.

Challenges and Findings in Crystallization

• The majority of crystals formed (over 95%) were highly twinned and had poor diffraction quality, only up to approximately 7-5 A resolution.
• However, diffraction-quality trigonal crystals were produced under certain conditions – a = b = 128.4, c = 85.8 A, and belonging to space group P3(2)21.
• A 2.9 A native data set was collected, with an R(merge) of 12.7%.
• Preliminary analyses provide a structural understanding for the specific roles of DDAO in the crystallization process of the proteolyzed form of PLRP2.